Patent application title: Human Growth Gene and Short Stature Gene Region

Inventors: Gudrun Rappold-Hoerbrand (Heidelberg, DE) Ercole Rao (Riedstadt, DE)
IPC8 Class: AA61K3827FI
USPC Class: 514 12
Class name: 25 or more peptide repeating units in known peptide chain structure
Publication date: 04/30/2009
Patent application number: 20090111744

Abstract:

Subject of the present invention is an isolated human nucleic acid molecule encoding polypeptides containing a homeobox domain of sixty amino acids having the amino acid sequence of SEQ ID NO: 1 and having regulating activity on human growth. Three novel genes residing within the about 500 kb short stature critical region on the X and Y chromosome were identified. At least one of these genes is responsible for the short stature phenotype. The cDNA corresponding to this gene may be used in diagnostic tools, and to further characterize the molecular basis for the short stature-phenotype. In addition, the identification of the gene product of the gene provides new means and methods for the development of superior therapies for short stature.

Claims:

1. A method for the treatment of short stature in patients identified as having a genetic: defect in the human growth gene SHOX, comprisinga) identifying a genetic defect in a human subject suspected of having a genetic mutation in the SHOX gene having the nucleotide sequence according to SEQ. ID NO. 14, andb) administering to said patient a therapeutically active amount of human growth hormone.

2. The method according to claim 1, wherein said genetic defect is identified by obtaining a biological sample molecule to be examined, and amplifying said biological sample molecule in the presence of two nucleotide probes completely or in part complementary to any of the DNA sequences of SEQ. ID. NO: 2 to SEQ ID NO. 7.

3. The method according to claim 1, wherein the genetic mutation is caused by a hot spot of mutation in the nucleic acid sequence encoding a protein truncation at amino acid position 195 in the SHOX gene.

4. The method according to claim 1, wherein said patient is not suffering from Turner's Syndrome.

5. A method for the treatment of short stature in patients identified as having a genetic defect in the human growth gene SHOX, said SHOX gene having the nucleotide sequence according to SEQ. ID NO. 14, comprisinga) identifying a genetic defect in a human subject suspected of having a genetic mutation in the SHOX gene, comprisingi) obtaining a biological sample containing a polynucleotide from a human subject,ii) amplifying the polynucleotide of i) in the presence of a primer, wherein said primer is an exon flanking primer or a primer to an exon nucleotide sequence of the SHOX gene according to SEQ ID NO: 14 and wherein the oligonucleotide primer has a length of 18-26 nucleotides and the oligonucleotide sequence of said primer is identical to a partial sequence of an exon nucleotide sequence of the SHOX gene according to SEQ ID NO: 14, andiii) sequencing any amplification product of the polynucleotide ofi) to determine the presence of a genetic mutation in the SHOX gene of said human subject, andb) administering to said human subject a therapeutically active amount of human growth hormone.

6. The method according to claim 5, wherein the exon nucleotide sequence in step a) ii) is a polynucleotide sequence selected from the group consisting of SHOX ET93 (SEQ ID NO: 2), SHOX G310 (SEQ ID NO: 3), SHOX ET45 (SEQ ID NO: 4), SHOX G108 (SEQ ID NO: 5), SHOX Va (SEQ ID NO: 6) and SHOX Vb (SEQ ID NO: 7).

Description:

[0001]This application is a divisional of U.S. Ser. No. 10/158,160 filed May 31, 2002, which is a continuation of U.S. Ser. No. 09/147,699, filed Jun. 24, 1999, now abandoned, which is a 371 of PCT/EP97/05355, filed on Sep. 29, 1997, which claims the benefit of U.S. provisional application 60/027,633, filed on Oct. 1, 1996.

[0002]The present invention relates to the isolation, identification and characterization of newly identified human genes responsible for disorders relating to human growth, especially for short stature or Turner syndrome, as well as the diagnosis and therapy of such disorders.

[0003]The isolated genomic DNA or fragments thereof can be used for pharmaceutical purposes or as diagnostic tools or reagents for identification or characterization of the genetic defect involved in such disorders. Subject of the present invention are further human growth proteins (transcription factors A, B and C) which are expressed after transcription of said DNA into RNA or mRNA and which can be used in the therapeutic treatment of disorders related to mutations in said genes. The invention further relates to appropriate cDNA sequences which can be used for the preparation of recombinant proteins suitable for the treatment of such disorders. Subject of the invention are further plasmid vectors for the expression of the DNA of these genes and appropriate cells containing such DNAs. It is a further subject of the present invention to provide means and methods for the genetic treatment of such disorders in the area of molecular medicine using an expression plasmid prepared by incorporating the DNA of this invention downstream from an expression promotor which effects expression in a mammalian host cell.

[0004]Growth is one of the fundamental aspects in the development of an organism, regulated by a highly organised and complex system. Height is a multifactorial trait, influenced by both environmental and genetic factors. Developmental malformations concerning body height are common phenomena among humans of all races. With an incidence of 3 in 100, growth retardation resulting in short stature account for the large majority of inborn deficiencies seen in humans.

[0005]With an incidence of 1:2500 life-born phenotypic females, Turner syndrome is a common chromosomal disorder (Rosenfeld et al., 1996). It has been estimated that 1-2% of all human conceptions are 45,X and that as many as 99% of such fetuses do not come to term (Hall and Gilchrist, 1990; Robins, 1990). Significant clinical variability exists in the phenotype of persons with Turner syndrome (or Ullrich-Turner syndrome) (Ullrich, 1930; Turner, 1938). Short stature, however, is a consistent finding and together with gonadal dysgenesis considered as the lead symptoms of this disorder. Turner syndrome is a true multifactorial disorder. Both the embryonic lethality, the short stature, gonadal dysgenesis and the characteristic somatic features are thought to be due to monosomy of genes common to the X and Y chromosomes. The diploid dosis of those X-Y homologous genes are suggested to be requested for normal human development. Turner genes (or anti-Turner genes) are expected to be expressed in females from both the active and inactive X chromosomes or Y chromosome to ensure correct dosage of gene product. Haploinsufficiency (deficiency due to only one active copy), consequently would be the suggested genetic mechanism underlying the disease.

[0006]A variety of mechanisms underlying short stature have been elucidated so far. Growth hormone and growth hormone receptor deficiencies as well as skeletal disorders have been described as causes for the short stature phenotype (Martial et al., 1979; Phillips et al., 1981; Leung et al., 1987; Goddard et al., 1995). Recently, mutations in three human fibroblast growth factor receptor-encoding genes (FGFR 1-3) were identified as the cause of various skeletal disorders, including the most common form of dwarfism, achondroplasia (Shiang et al., 1994; Rousseau et al., 1994; Muenke and Schell, 1995). A well-known and frequent (1:2500 females) chromosomal disorder, Turner Syndrome (45,X), is also consistently associated with short stature. Taken together, however, all these different known causes account for only a small fraction of all short patients, leaving the vast majority of short stature cases unexplained to date.

[0007]The sex chromosomes X and Y are believed to harbor genes influencing height (Ogata and Matsuo, 1993). This could be deduced from genotype-phenotype correlations in patients with sex chromosome abnormalities. Cytogenetic studies have provided evidence that terminal deletions of the short arms of either the X or the Y chromosome consistently lead to short stature in the respective individuals (Zuffardi et al., 1982; Curry et al., 1984). More than 20 chromosomal rearrangements associated with terminal deletions of chromosome Xp and Yp have been reported that localize the gene(s) responsible for short stature to the pseudoautosomal region (PAR1) (Ballabio et al., 1989, Schaefer et al., 1993). This localisation has been narrowed down to the most distal 700 kb of DNA of the PAR1 region, with DXYS 15 as the flanking marker (Ogata et al., 1992; 1995).

[0008]Mammalian growth regulation is organized as a complex system. It is conceivable that multiple growth promoting genes (proteins) interact with one another in a highly organized way. One of those genes controlling height has tentatively been mapped to the pseudoautosomal region PAR1 (Ballabio et al., 1989), a region known to be freely exchanged between the X and Y chromosomes (for a review see Rappold, 1993). The entire PAR1 region is approximately 2,700 kb.

[0009]The critical region for short stature has been defined with deletion patients. Short stature is the consequence when an entire 700 kb region is deleted or when a specific gene within this critical region is present in haploid state, is interrupted or mutated (as is the case with idiotypic short stature or Turner sydrome). The frequency of Turner's syndrome is 1 in 2500 females worldwide; the frequency of this kind of idiopathic short stature can be estimated to be 1 in 4.000-5.000 persons. Turner females and some short stature individuals usually receive an unspecific treatment with growth hormone (GH) for many years to over a decade although it is well known that they have normal GH levels and GH deficiency is not the problem. The treatment of such patients is very expensive (estimated costs approximately 30.000 USD p.a.). Therefore, the problem existed to provide a method and means for distinguishing short stature patients on the one side who have a genetic defect in the respective gene and on the other side patients who do not have any genetic defect in this gene. Patients with a genetic defect in the respective gene--either a complete gene deletion (as in Turner syndrome) or a point mutation (as in idiopathic short stature)--should be susceptible for an alternative treatment without human GH, which now can be devised.

[0010]Genotype/phenotype correlations have supported the existence of a growth gene in the proximal part of Yq and in the distal part of Yp. Short stature is also consistently found in individuals with terminal deletions of Xp. Recently, an extensive search for male and female patients with partial monosomies of the pseudoautosomal region has been undertaken. On the basis of genotype-phenotype correlations, a minimal common region of deletion of 700 kb DNA adjacent to the telomere was determined (Ogata et al., 1992; Ogata et al., 1995). The region of interest was shown to lie between genetic markers DXYS20 (3cosPP) and DXYS15 (113D) and all candidate genes for growth control from within the PAR1 region (e.g., the hemopoietic growth factor receptor a; CSF2RA) (Gough et al., 1990) were excluded based on their physical location (Rappold et al., 1992). That is, the genes were within the 700 kb deletion region of the 2.700 kb PAR1 region.

[0011]Deletions of the pseudoautosomal region (PAR1) of the sex chromosomes were recently discovered in individuals with short stature and subsequently a minimal common deletion region of 700 kb within PAR1 was defined. Southern blot analysis on DNA of patients AK and SS using different pseudoautosomal markers has identified an Xp terminal deletion of about 700 kb distal to DXYS 15 (113D) (Ogata et al, 1992; Ogata et al, 1995).

[0012]The gene region corresponding to short stature has been identified as a region of approximately 500 kb, preferably approximately 170 kb in the PAR1 region of the X and Y chromosomes. Three genes in this region have been identified as candidates for the short stature gene. These genes were designated SHOX (also referred to as SHOX93 or HOX93), (SHOX=short stature homeobox-containing gene), pET92 and SHOT (SHOX-like homeobox gene on chromosome three). The gene SHOX which has two separate splicing sites resulting in two variations (SHOX a and b) is of particular importance. In preliminary investigations, essential parts of the nucleotide sequence of the short stature gene could be analysed (SEQ ID No. 8). Respective exons or parts thereof could be predicted and identified (e.g. exon I [G310]; exon II [ET93]; exon IV [G108]; pET92). The obtained sequence information could then be used for designing appropriate primers or nucleotide probes which hybridize to parts of the SHOX gene or fragments thereof. By conventional methods, the SHOX gene can then be isolated. By further analysis of the DNA sequence of the genes responsible for short stature, the nucleotide sequence of exons I-V could be refined (v. FIG. 1-3). The gene SHOX contains a homeobox sequence (SEQ ID NO: I) of approximately 180 bp (v. FIG. 2 and FIG. 3), starting from the nucleotide coding for amino acid position 117 (Q) to the nucleotide coding for amino acid position 176 (E), i.e. from CAG (440) to GAG (619). The homeobox sequence is identified as the homeobox-pET93 (SHOX) sequence and two point mutations have been found in individuals with short stature in a German (A1) and a Japanese patient by screening up to date 250 individuals with idiopahtic short stature. Both point mutations were found at the identical position and leading to a protein truncation at amino acid position 195, suggesting that there may exist a hot spot of mutation. Due to the fact that both mutations found, which lead to a protein truncation, are at the identical position, it is possible that a putative hot spot of recombination exisits with exon 4 (G108). Exon specific primers can therefore be used as indicated below, e.g. GCA CAG CCA ACC ACC TAG (for) or TGG AAA GGC ATC ATC CGT AAG (rev).

[0013]The above-mentioned novel homeobox-containing gene, SHOX, which is located within the 170 kb interval, is alternatively spliced generating two proteins with diverse function. Mutation analysis and DNA sequencing were used to demonstrate that short stature can be caused by mutations in SHOX.

[0014]The identification and cloning of the short stature critical region according to the present invention was performed as follows: Extensive physical mapping studies on 15 individuals with partial monosomy in the pseudoautosomal region (PAR1) were performed. By correlating the height of those individuals with their deletion breakpoints a short stature (SS) critical region of approximately 700 kb was defined. This region was subsequently cloned as an overlapping cosmid contig using yeast artificial chromosomes (YACs) from PAR 1 (Ried et al., 1996) and by cosmid walking. To search for candidate genes for SS within this interval, a variety of techniques were applied to an approximately 600 kb region between the distal end of cosmid 56G10 and the proximal end of 51D11. Using cDNA selection, exon trapping, and CpG island cloning, the two novel genes were identified.

[0015]The position of the short stature critical interval could be refined to a smaller interval of 170 kb of DNA by characterizing three further specific individuals (GA, AT and RY), who were consistently short. To precisely localize the rearrangement breakpoints of those individuals, fluorescence in situ hybridization (FISH) on metaphase chromosomes was carried out using cosmids from the contig. Patient GA, with a terminal deletion and normal height, defined the distal boundary of the critical region (with the breakpoint on cosmid 110E3), and patient AT, with an X chromosome inversion and normal height, the proximal boundary (with the breakpoint on cosmid 34F5). The Y-chromosomal breakpoint of patient RY, with a terminal deletion and short stature, was also found to be contained on cosmid 34F5, suggesting that this region contains sequences predisposing to chromosome rearrangements.

[0016]The entire region, bounded by the Xp/Yp telomere, has been cloned as a set of overlapping cosmids. Fluorescence in situ hybridization (FISH) with cosmids from this region was used to study six patients with X chromosomal rearrangements, three with normal height and three with short stature. Genotype-phenotype correlations narrowed down the critical short stature interval to 270 kb of DNA or even less as 170 kb, containing the gene or genes with an important role in human growth. A minimal tiling path of six to eight cosmids bridging this interval is now available for interphase and metaphase FISH providing a valuable tool for diagnostic investigations on patients with idiopathic short stature.

BRIEF DESCRIPTION OF THE DRAWINGS

[0017]FIG. 1 is a gene map of the SHOX gene including five exons which are identified as follows: exon I: G310, exon II: ET93, exon III: ET45, exon IV: G108 and exons Va and Vb, whereby exons Va and Vb result from two different splicing sites of the SHOX gene. Exon II and III contain the homeobox sequence of 180 nucleotides.

[0018]FIGS. 2 and 3 are the nucleotide and predicted amino acid sequences of SHOXa and SHOXb: [0019]SHOX a: The predicted start of translation begins at nucleotide 92 with the first in-frame stop codon (TGA) at nucleotides 968-970, yielding an open reading frame of 876 bp that encodes a predicted protein of 292 amino acids (designated as transcription factor A or SHOXa protein, respectively). An in-frame, 5'stop codon at nucleotide 4, the start codon and the predicted termination stop codon are in bold. The homeobox is boxed (starting from amino acid position 117 (Q) to 176 (E), i.e. CAG thru GAG in the nucleotide sequence). The locations of introns are indicated with arrows. Two putative polyadenylation signals in the 3'untranslated region are underlined. [0020]SHOX b: An open reading frame of 876 bp exists from A in the first methionin at nucleotide 92 to the in-frame stop codon at nucleotide 767-769, yielding an open reading frame of 675 bp that encodes a predicted protein of 225 amino acids (transcription factor B or SHOXb protein, respectively). The locations of introns are indicated with arrows. Exons I-IV are identical with SHOXa, exon V is specific for SHOX b. A putative polyadenylation signal in the 3' untranslated region is underlined.

[0021]FIG. 4 are the nucleotide (SEQ ID NO:43) and predicted amino acid (SEQ ID NO: 16) sequence of SHOT. The predicted start of translation begins at nucleotide 43 with the first in-frame stop codon (TGA) at nucleotides 613-615, yielding an open reading frame of 573 bp that encodes a predicted protein of 190 amino acids (designated as transcription factor C or SHOT protein, respectively). The homeobox is boxed (starting from amino acid position 11 (Q) to 70 (E), i.e. CAG thru GAG in the nucleotide sequence). The locations of introns are indicated with arrows. Two putative polyadenylation signals in the 3'untranslated region are underlined

[0022]FIG. 5 gives the exon/intron organization of the human SHOX gene and the respective positions in the nucleotide sequence (Intron/Exon sequences (SEQ ID NOS 44-49, respectively in order of appearance) and Exon/Intron sequences (SEQ ID NOS 50-55, respectively in order of appearance)).

BRIEF DESCRIPTION OF THE SEQ ID:

[0023]SEQ ID NO.1: translated amino acid sequence of the homeobox domain (180 bp)SEQ ID NO.2: exon II (ET93) of the SHOX geneSEQ ID NO. 3: exon I (G310) of the SHOX geneSEQ ID NO. 4: exon III (ET45) of the SHOX geneSEQ ID NO. 5: exon IV (G108) of the SHOX geneSEQ ID NO. 6: exon Va of the SHOX geneSEQ ID NO. 7: exon Vb of the SHOX geneSEQ ID NO. 8: preliminary nucleotide sequence of the SHOX geneSEQ ID NO.9: ET92 geneSEQ ID NO.10: SHOXa sequence (see also FIG. 2)SEQ ID NO.1: transcription factor A (see also FIG. 2)SEQ ID NO. 12: SHOXb sequence (see also FIG. 3)SEQ ID NO. 13: transcription factor B (see also FIG. 3)SEQ ID NO. 14: SHOX geneSEQ ID NO. 15: SHOT sequenceSEQ ID NO. 16: transcription factor C (see also FIG. 4)

[0024]Since the target gene leading to disorders in human growth (e.g. short stature region) was unknown prior to the present invention, the biological and clinical association of patients with this deletion could give insights to the function of this gene. In the present study, fluorescence in situ hybridization (FISH) was used to examine metaphase and interphase lymphocyte nuclei of six patients. The aim was to test all cosmids of the overlapping set for their utility as FISH probes and to determine the breakpoint regions in all four cases, thereby determining the minimal critical region for the short stature gene.

[0025]Duplication and deletion of genomic DNA can be technically assessed by carefully controlled quantitative PCR or dose estimation on Southern blots or by using RFLPs. However, a particularly reliable method for the accurate distinction between single and double dose of markers is FISH, the clinical application of is presently routine. Whereas in interphase FISH, the pure absence or presence of a molecular marker can be evaluated, FISH on metaphase chromosomes may provide a semi-quantitative measurement of inter-cosmid deletions. The present inventor has determined that deletions of about 10 kb (25% of signal reduction) can still be detected. This is of importance, as practically all disease genes on the human X chromosome have been associated with smaller and larger deletions in the range from a few kilobases to several megabases of DNA (Nelson et al., 1995).

[0026]Subject of the present invention are therefore DNA sequences or fragments thereof which are part of the genes responsible for human growth (or for short stature, respectively, in case of genetic defects in these genes). Three genes responsible for human growth were identified: SHOX, pET92 and SHOT. DNA sequences or fragments of these genes, as well as the respective full length DNA sequences of these genes can be transformed in an appropriate vector and transfected into cells. When such vectors are introduced into cells in an appropriate way as they are present in healthy humans, it is devisable to treat diseases involved with short stature, i.e. Turners syndrome, by modern means of gene therapy. For example, short stature can be treated by removing the respective mutated growth genes responsible for short stature. It is also possible to stimulate the respective genes which compensate the action of the genes responsible for short stature, i.e. by inserting DNA sequences before, after or within the growth/short stature genes in order to increase the expression of the healthy allels. By such modifications of the genes, the growth/short stature genes become activated or silent, respectively. This can be accomplished by inserting DNA sequences at appropriate sites within or adjacent to the gene, so that these inserted DNA sequences interfere with the growth/short stature genes and thereby activate or prevent their transcription. It is also devisable to insert a regulatory element (e.g. a promotor sequence) before said growth genes to stimulate the genes to become active. It is further devisable to stimulate the respective promotor sequence in order to overexpress--in the case of Turner syndrome--the healthy functional allele and to compensate for the missing allele. The modification of genes can be generally achieved by inserting exogenous DNA sequences into the growth gene/short stature gene via homologous recombination.

[0027]The DNA sequences according to the present invention can also be used for transformation of said sequences into animals, such as mammals, via an appropriate vector system. These transgenic animals can then be used for in vivo investigations for screening or identifying pharamceutical agents which are useful in the treatment of diseases involved with short stature. If the animals positively respond to the administration of a candidate compound or agent, such agent or compound or derivatives thereof would be devisable as pharmaceutical agents. By appropriate means, the DNA sequences of the present invention can also be used in genetic experiments aiming at finding methods in order to compensate for the loss of genes responsible for short stature (knock-out animals).

[0028]In a further object of this invention, the DNA sequences can also be used to be transformed into cells. These cells can be used for identifying pharmaceutical agents useful for the treatment of diseases involved with short stature, or for screening of such compounds or library of compounds. In an appropriate test system, variations in the phenotype or in the expression pattern of these cells can be determined, thereby allowing the identification of interesting candidate agents in the development of pharmaceutical drugs.

[0029]The DNA sequences of the present invention can also be used for the design of appropriate primers which hybridize with segments of the short stature genes or fragments thereof under stringent conditions. Appropriate primer sequences can be constructed which are useful in the diagnosis of people who have a genetic defect causing short stature. In this respect it is noteworthy that the two mutations found occur at the identical position, suggesting that a mutational hot spot exists.

[0030]In general, DNA sequences according to the present invention are understood to embrace also such DNA sequences which are degenerate to the specific sequences shown, based on the degeneracy of the genetic code, or which hybridize under stringent conditions with the specifically shown DNA sequences.

[0031]The present invention encompasses especially the following aspects: [0032]a) An isolated human nucleic acid molecule encoding polypeptides containing a homeobox domain of sixty amino acids having the amino acid sequence of SEQ ID NO: 1 and having regulating activity on human growth. [0033]b) An isolated DNA molecule comprising the nucleotide sequence essentially as indicated in FIG. 2, FIG. 3 or FIG. 4, and especially as shown in SEQ ID NO: 10, SEQ ID NO: 12 or SEQ ID NO: 15. [0034]c) DNA molecules capable of hybridizing to the DNA molecules of item b). [0035]d) DNA molecules of item c) above which are capable of hybridization with the DNA molecules of item 2. under a temperature of 60-70° C. and in the presence of a standard buffer solution. [0036]e) DNA molecules comprising a nucleotide sequence having a homology of seventy percent or higher with the nucleotide sequence of SEQ ID NO: 10, SEQ ID NO: 12 or SEQ ID NO: 15 and encoding a polypeptide having regulating activity on human growth. [0037]f) Human growth proteins having the amino acid sequence of SEQ ID NO: 11, 13 or 16 or a functional fragment thereof. [0038]g) Antibodies obtained from immunization of animals with human growth proteins of item f) or antigenic variants thereof. [0039]h) Pharmaceutical compositions comprising human growth proteins or functional fragments thereof for treating disorders caused by genetic mutations of the human growth gene. [0040]i) A method of screening for a substance effective for the treatment of disorders mentioned above under item h) comprising detecting messenger RNA hybridizing to any of the DNA molecules decribed in a)-e) so as to measure any enhancement in the expression levels of the DNA molecule in response to treatment of the host cell with that substance. [0041]j) An expression vector or plasmid containing any of the nucleic acid molecules described in a)-e) above which enables the DNA molecules to be expressed in mammalian cells. [0042]k) A method for the determination of the gene or genes responsible for short stature in a biological sample of body tissues or body fluids.

[0043]In the method k) above, preferably nucleotide amplification techniques, e.g. PCR, are used for detecting specific nucleotide sequences known to persons skilled in the art, and described, for example, by Mullis et al. 1986, Cold Spring Harbor Symposium Quant. Biol. 51, 263-273, and Saiki et al., 1988, Science 239, 487-491, which are incorporated herein by reference. The short stature nucleotide sequences to be determined are mainly those represented by sequences SEQ ID No. 2 to SEQ ID No. 7.

[0044]In principle, all oligonucleotide primers and probes for amplifying and detecting a genetic defect responsible for deminished human growth in a biological sample are suitable for amplifying a target short stature associated sequence. Especially, suitable exon specific primer pairs according to the invention are provided by table 1. Subsequently, a suitable detection, e.g. a radioactive or non-radioactive label is carried out.

TABLE-US-00001 TABLE 1 Exon Sense primer Antisense primer Product (bp) Ta (° C.) 5'-I (G310) SP 1 ASP 1 194 58 3'-I (G310) SP 2 ASP 2 295 58 II (ET93) SP 3 ASP 3 262 76/72/68 III (ET45) SP 4 ASP 4 120 65 IV (G108) SP 5 ASP 5 154 62 Va (SHOXa) SP 6 ASP 6 265 61

explanation of the abbreviations for the primers:

TABLE-US-00002 SP1 ATTTCCAATGGAAAGGCGTAAATAAC SP2 ACGGCTTTTGTATCCAAGTCTTTTG SP3 GCCCTGTGCCCTCCGCTCCC SP4 GGCTCTTCACATCTCTCTCTGCTTC SP5 CCACACTGACACCTGCTCCCTTTG S5P6 CCCGCAGGTCCAGGCTCAGCTG ASP1 CGCCTCCGCCGTTACCGTCCTTG ASP2 CCCTGGAGCCGGCGCGCAAAG ASP3 CCCCGCCCCCGCCCCCGG ASP4 CTTCAGGTCCCCCCAGTCCCG ASP5 CTAGGGATCTTCAGAGGAAGAAAAAG ASP6 GCTGCGCGGCGGGTCAGAGCCCCAG

[0045]Also, a single stranded RNA can be used as target. Methods for reversed transcribing RNA into cDNA are also well known and described in Sambrook et al., Molecular Cloning: A Laboratory Manual, New York, Cold Spring Harbor Laboratory 1989. Alternatively, preferred methods for reversed transcription utilize thermostable DNA polymerases having RT activity.

[0046]Further, the technique described before can be used for selecting those person from a group of persons being of short stature characterized by a genetic defect and which allows as a consequence a more specific medical treatment.

[0047]In another subject of the present invention, the transcription factors A, B and C can be used as pharmaceutical agents. These transcription factors initiate a still unknown cascade of biological effects on a molecular level involved with human growth. These proteins or functional fragments thereof have a mitogenic effect on various cells. Especially, they have an osteogenic effect. They can be used in the treatment of bone diseases, such as e.g. osteoporosis, and especially all those diseases involved with disturbance in the bone calcium regulation.

[0048]As used herein, the term "isolated" refers to the original derivation of the DNA molecule by cloning. It is to be understood however, that this term is not intended to be so limiting and, in fact, the present invention relates to both naturally occurring and synthetically prepared seqences, as will be understood by the skilled person in the art.

[0049]The DNA molecules of this invention may be used in forms of gene therapy involving the use of an expression plasmid prepared by incorporating an appropriate DNA sequence of this invention downstream from an expression promotor that effects expression in a mammalian host cell. Suitable host cells are procaryotic or eucaryotic cells. Procaryotic host cells are, for example, E. coli, Bacillus subtilis, and the like. By transfecting host cells with replicons originating from species adaptable to the host, that is, plasmid vectors containing replication starting point and regulator sequences, these host cells can be transfected with the desired gene or cDNA. Such vectors are preferably those having a sequence that provides the transfected cells with a property (phenotype) by which they can be selected. For example, for E. coli hosts the strain E. coli K12 is typically used, and for the vector either pBR322 or pUC plasmids can be generally employed. Examples for suitable promoters for E. coli hosts are trp promotor, lac promotor or Ipp promotor. If desired, secretion of the expression product through the cell membrane can be effected by connecting a DNA sequence coding for a signal peptide sequence at the 5' upstream side of the gene. Eucaryotic host cells include cells derived from vertebrates or yeast etc.. As a vertebrate host cell, COS cells can be used (Cell, 1981, 23: 175-182), or CHO cells. Preferably, promotors can be used which are positioned 5' upstream of the gene to be expressed and having RNA splicing positions, polyadenylation and transcription termination seqences.

[0050]The transcription factors A, B and C of the present invention can be used to treat disorders caused by mutations in the human growth genes and can be used as growth promoting agents. Due to the polymorphism known in the case of eukaryotic genes, one or more amino acids may be substituted. Also, one or more amino acids in the polypeptides can be deleted or inserted at one or more sites in the amino acid sequence of the polypeptides of SEQ ID NO: 11, 13 or 16. Such polypeptides are generally referred to equivalent polypeptides as long as the underlying biological acitivity of the unmodified polypeptide remains essentially unchanged.

[0051]The present invention is illustrated by the following examples.

EXAMPLE 1

Patients

[0052]All six patients studied had de novo sex chromosome aberrations.

[0053]CC is a girl with a karyotype 45,X/46,X psu dic (X) (Xqter→Xp22.3::Xp22.3→Xqter). At the last examination at 61/2 years of age, her height was 114 cm (25-50 the % percentile). Her mother's height was 155 cm, the father was not available for analysis. For details, see Henke et al., 1991.

[0054]GA is a girl with a karyotype 46,X der X (3pter→3p23::Xp22.3→Xqter). At the last examination at 17 years, normal stature (159 cm) was observed. Her mother's height is 160 cm and her father's height 182 cm. For details, see Kulharya et al, 1995.

[0055]SS is a girl with a karyotype 46,X rea (X) (Xqter→Xq26::Xp22.3→Xq26:). At 11 years her height remained below the 3rd percentile growth curve for Japanese girls; her predicted adult height (148.5 cm) was below her target height (163 cm) and target range (155 to 191 em). For details, see Ogata et alt, 1992.

[0056]AK is a girl with a karyotype 46,X rea (X) (Xqter→Xp22.3::Xp22.3→Xp21.3:). At 13 years her height remained below the 2nd percentile growth curve for Japanese girls; her predicted adult height (142.8 cm) was below her target height (155.5 cm) and target range (147.5-163.5 em). For details, see Ogata et alt, 1995.

[0057]RY: the karyotype of the ring Y patient is 46,X,r(Y)/46,Xdic r(Y)/45,X[95:3:2], as examined on 100 lymphocytes; at 16 years of age his final height was 148; the heights of his three brothers are all in the normal range with 170 cm (16 years, brother 1), 164 cm (14 years, brother 2) and 128 cm (9 years, brother 3), respectively. Growth retardation of this patient is so severe that it would also be compatible with an additional deletion of the GCY locus on Yq.

[0058]AT: boy with ataxia and inv(X); normal height of 116 cm at age 7, parents' heights are 156 cm and 190 cm, respectively.

Patients for Mutation Analysis:

[0059]250 individuals with idiopathic short stature were tested for mutations in SHOXa. The patients were selected on the following criteria: height for chronological age was below the 3rd centile of national height standards, minus 2 standard deviations (SDS); no causative disease was known, in particular: normal weight (length) for gestational age, normal body proportions, no chronic organic disorder, normal food intake, no psychiatric disorder, no skeletal dysplasia disorder, no thyroid or growth hormone deficiency.

Family A:

[0060]Cases 1 and 2 are short statured children of a German non-consanguineous family. The boy (case 1) was born at the 38th week of gestation by cesarian section. Birth weight was 2660 g, birth length 47 cm. He developed normally except for subnormal growth. On examination at the age of 6.4 years, he was proportionate small (106.8 cm, -2.6 SDS) and obese (22.7 kg), but otherwise normal. His bone age was not retarded (6 yrs) and bone dysplasia was excluded by X-ray analysis. IGF-I and IGFBP-3 levels as well as thyroid parameters in serum rendered GH or thyroid hormone deficiency unlikely. The girl (case 2) was born at term by cesarian section. Birth weight was 2920 g, birth length 47 cm. Her developmental milestones were normal, but by the age of 12 months poor growth was apparent (length: 67 cm, -3.0 SDS). At 4 years she was 89.6 cm of height (-3.6 SDS). No dysmorphic features or dysproportions were apparent. She was not obese (13 kg). Her bone age was 3.5 years and bone dysplasia was excluded. Hormone parameters were normal. It is interesting to note that both the girl and the boy grow on the 50 percentile growth curve for females with Turner syndrome. The mother is the smallest of the family and has a mild rhizomelic dysproportion (142.3 cm, -3.8 SDS). One of her two sisters (150 cm, -2.5 SDS) and the maternal grandmother (153 cm, -2.0 SDS) are all short without any dysproportion. One sister has normal stature (167 cm, +0.4 SDS). The father's height is 166 cm (-1.8 SDS) and the maternal grandfather' height is 165 cm (-1.9 SDS). The other patient was of Japanese origin and showed the identical mutation.

EXAMPLE 2

Identification of the Short Stature Gene

[0061]A. In situ Hybridizationa) Florescence in situ Hybridization (FISH)

[0062]Florescence in situ hybridization (FISH) using cosmids residing in the Xp/Yp pseudoautosomal region (PAR1) was carried out. FISH studies using cosmids 64/75cos (LLNLc 110H032), E22cos (2e2), F1/14cos (110A7), M1/70cos (110E3), P99F2cos (43C11), P99cos (LLNLc110P2410), B6cosb (1CRFc104H0425), F20cos (34F5), F21cos (ICRFc104G0411), F3cos2 (9E3), F3cos1 (11E6), P117cos (29B11), P6cos1 (ICRFc104P0117), P6cos2 (LLNLc110E0625) and E4cos (15G7) was carried out according to published methods (Lichter and Cremer, 1992). In short, one microgram of the respective cosmid clone was labeled with biotin and hybridized to human metaphase chromosomes under conditions that suppress signals from repetitive DNA sequences. Detection of the hybridization signal was via FITC-conjugated avidin. Images of FITC were taken by using a cooled charge coupled device camera system (Photometrics, Tucson, Ariz.).

b) Physical Mapping

[0063]Cosmids were derived from Lawrence Livermore National Laboratory X- and Y-chromosome libraries and the Imperial Cancer Research Fund London (now Max Planck Institute for Molecular Genetics Berlin) X chromosome library. Using cosmids distal to DXYS15, namely E4cos, P6cos2, P6cos1, P1117cos and F3cos1 one can determine that two copies are still present of E4cos, P6cos2, P6cos1 and one copy of P117cos and F3cos1. Breakpoints of both patients AK and SS map on cosmid P6cos1, with a maximum physical distance of 10 kb from each other. It was concluded that the abnormal X chromosomes of AK and SS have deleted about 630 kb of DNA. Further cosmids were derived from the ICRF X chromosome specific cosmid library (ICRFc104), the Lawrence Livermore X chromosome specific cosmid library (LLNLc110) and the Y chromosome specific library (LLCO3'M'), as well as from a self-made cosmid library covering the entire genome. Cosmids were identified by hybridisation with all known probes mapping to this region and by using entire YACs as probes. To verify overlaps, end probes from several cosmids were used in cases in which overlaps could not be proven using known probes.

c) Southern Blot Hybridisation

[0064]Southern blot analysis using different pseudoautosomal markers has provided evidence that the breakpoint on the X chromosome of patient CC resides between DXYS20 (3cosPP) and DXYS60 (U7A) (Henke et al, 1991). In order to confirm this finding and to refine the breakpoint location, cosmids 64/75cos, E22cos, F1/14cos, M1/70cos, F2cos, P99F2cos and P99cos were used as FISH probes. The breakpoint location on the abnormal X of patient CC between cosmids 64/75cos (one copy) and F1/14cos (two copies) on the E22PAC could be determined. Patient CC with normal stature consequently has lost approximately 260-290 kb of DNA.

[0065]Southern blot hybridisations were carried out at high stringency conditions in Church buffer (0.5 M NaPi pH 7.2, 7% SDS, 1 mM EDTA) at 65° C. and washed in 40 mM NaPi, 1% SDS at 65° C.

d) FISH Analysis

[0066]Biotinylated cosmid DNA (insert size 32-45 kb) or cosmid fragments (10-16 kb) were hybridised to metaphase chromosomes from stimulated lymphocytes of patients under conditions as described previously (Lichter and Cremer, 1992). The hybridised probe was detected via avidin-conjugated FITC.

e) PCR Amplification

[0067]All PCRs were performed in 50 μl volumes containing 100 pg-200 ng template, 20 pmol of each primer, 200 μM dNTP's (Pharmacia), 1.5 mM MgCl₂, 75 mMTris/HCl pH9, 20 mM (NH₄)₂SO₄ 0.01% (w/v) Tween20 and 2 U of Goldstar DNA Polymerase (Eurogentec). Thermal cycling was carried out in a Thermocycler GeneE (Techne).

f) Exon Amplification

[0068]Four cosmid pools consisting of each four to five clones from the cosmid contigs were used for exon amplification experiments. The cosmids in each cosmid pool were partially digested with Sau3A. Gel purified fractions in the size range of 4-10 kb were cloned in the BamHI digested pSPL3B vector (Burn et al, 1995) and used for the exon amplification experiments as previously described (Church et al., 1994).

g) Genomic Sequencing

[0069]Sonificated fragments of the two cosmids LLOYNCO3'M'15D10 and LLOYNCO3'M'34F5 were subcloned separately into M13mp18 vectors. From each cosmid library at least 1000 plaques were picked, M13 DNA prepared and sequenced using dye-terminators, ThermoSequenase (Amersham) and universal M13-primer (MWG-BioTech). The gels were run on ABI-377 sequencers and data were assembled and edited with the GAP4 program (Staden).

[0070]Of all six patients, GA had the least well characterized chromosomal breakpoint. The most distal markers previously tested for their presence or absence on the X were DXS1060 and DXS996, which map approximately 6 Mb from the telomere (Nelson et al., 1995). Several cosmids containing different gene sequences from within PAR1 (MIC2, ANT3, CSF2RA, and XE7) were tested and all were present on the translocation chromosome. Cosmids from within the short stature critical region e.g., chromosome, thereby placing the translocation breakpoint on cosmid M1/70cos. A quantitative comparison of the signal intensities of M1/70cos between the normal and the rearranged X indicates that approximately 70% of this cosmid is deleted.

TABLE-US-00003 TABLE 2 Table 2: This table summarizes the FISH data for the 16 cosmids tested on four patients. CC GA AK SS 64/75cos - - E22cos - - F1/14cos + - M1/70cos + (+) F2cos + + P99F2cos + + P99cos + + B6cos + F20cos F21cos F3cos2 F3cos1 - - P117cos - - P6cos1 + + P6cos2 + + E4cos + + [-] one copy; indicates that the respective cosmid was deleted on the rearranged X, but present on the normal X chromosome [+] two copies; indicates that the respective cosmid is present on the rearranged and on the normal X chromosome [(+)] breakpoint region; indicates that the breakpoint occurs within the cosmid as shown by FISH

[0071]In summary, the molecular analysis on six patients with X chromosomal rearrangements using florescence-labeled cosmid probes and in situ hybridization indicates that the short stature critical region can be narrowed down to a 270 kb interval, bounded by the breakpoint of patient GA from its centromere distal side and by patients AK and SS on its centromere proximal side.

[0072]Genotype-phenotype correlations may be informative and have been chosen to delineate the short stature critical interval on the human X and Y chromosome. In the present study FISH analysis was used to study metaphase spreads and interphase nuclei of lymphocytes from patients carrying deletions and translocations on the X chromosome and breakpoints within Xp22.3. These breakpoints appear to be clustered in two of the four patients (AK and SS) presumably due to the presence of sequences predisposing to chromosome rearrangements. One additional patient Ring Y has been found with an interruption in the 270 kb critical region, thereby reducing the critical interval to a 170 kb region.

[0073]By correlating the height of all six individuals with their deletion breakpoint, an interval of 170 kb was mapped to within the pseudoautosomal region, presence or absence of which has a significant effect on stature. This interval is bounded by the X chromosomal breakpoint of patient GA at 340 kb from the telomere (Xptel) distally and by the breakpoints of patients AT and RY at 510/520 kb Xptel proximally. This assignment constitutes a considerable reduction of the critical interval to almost one fourth of its previous size (Ogata et al., 1992; Ogata et al., 1995). A small set of six to eight cosmids are now available for FISH experiments to test for the prevalence and significance of this genomic locus on a large series of patients with idiopathic short stature.

B. Identification of the Candidate Short Stature Gene

[0074]To search for transcription units within the smallest 170 kb critical region, exon trapping and cDNA selection on six cosmids (110E3, F2cos, 43C11, P2410, 15D10, 34F5) was carried out. Three different positive clones (ET93, ET45 and G108) were isolated by exon trapping, all of which mapped back to cosmid 34F5. Previous studies using cDNA selection protocols and an excess of 25 different cDNA libraries had proven unsuccessful, suggesting that genes in this interval are expressed at very low abundancy.

[0075]To find out whether any gene in this interval was missed, the nucleotide sequence of about 140 kb from this region of the PAR1 was determined, using the random M13 method and dye terminator chemistry. The cosmids for sequence analysis were chosen to minimally overlap with each other and to collectively span the critical interval. DNA sequence analysis and subsequent protein prediction by the "X Grail" program, version 1.3c as well as by the exon-trapping program FEXHB were carried out and confirmed all 3 previously cloned exons. No protein-coding genes other than the previously isolated one could be detected.

C. Isolation of the Short Stature Candidate Gene SHOX

[0076]Assuming that all three exon clones ET93, ET45 and G108 are part of the same gene, they were used collectively as probes to screen 14 different cDNA libraries from 12 different fetal (lung, liver, brain 1 and 2) and adult tissues (ovary, placenta 1 and 2, fibroblast, skeletal muscle, bone marrow, brain, brain stem, hypothalamus, pituitary). Not a single clone among approximately 14 million plated clones was detected. To isolate the full-length transcript, 3' and 5'RACE were carried out. For 3'RACE, primers from exon G108 were used on RNA from placenta, skeletal muscle and bone marrow fibroblasts, tissues where G108 was shown to be expressed in. Two different 3'RACE clones of 1173 and 652 bp were derived from all three tissues, suggesting that two different 3'exons a and b exist. The two different forms were termed SHOXa and SHOXb.

[0077]To increase chances to isolate the complete 5'portion of a gene known to be expressed at low abundancy, a Hela cell line was treated with retinoic acid and phorbol ester PMA. RNA from such an induced cell line and RNA from placenta and skeletal muscle were used for the construction of a `Marathon cDNA library`. Identical 5'RACE cDNA clones were isolated from all three tissues.

Experimental Procedure:

[0078]RT-PCR and cDNA Library Construction

[0079]Human polyA⁺RNA of heart, pancreas, placenta, skeletal muscle, fetal kidney and liver was purchased from Clontech. Total RNA was isolated from a bone marrow fibroblast cell line with TRIZOL reagent (Gibco-BRL) as described by the manufacturer. First strand cDNA synthesis was performed with the Superscript first strand cDNA synthesis kit (Gibco-BRL) starting with 100 ng polyA⁺RNA or 10 μg total RNA using oligo(dt)-adapter primer (GGCCACGCGTCGACTAGTAC[dT]₂₀N. After first strand cDNA synthesis the reaction mix was diluted 1/10. For further PCR experiments 5 μl of this dilutions were used.

[0080]A `Marathon CDNA library` was constructed from skeletal muscle and placenta polyA⁺ RNA with the marathon cDNA amplification kit (Clontech) as described by the manufacturer.

[0081]Fetal brain (catalog # HL5015b), fetal lung (HL3022a), ovary (HL1098a), pituitary gland (HL1097v) and hypothalamus (HL1172b) cDNA libraries were purchased from Clontech. Brain, kidney, liver and lung cDNA libraries were part of the quick screen human cDNA library panel (Clontech). Fetal muscle cDNA library was obtained from the UK Human Genome Mapping Project Resource Center.

D. Sequence Analysis and Structure of SHOX Gene

[0082]A consensus sequence of SHOXa and SHOXb (1349 and 1870 bp) was assembled by analysis of sequences from the 5' and 3'RACE derived clones. A single open reading frame of 1870 bp (SHOXa) and 1349 bp (SHOXb) was identified, resulting in two proteins of 292 (SHOXa) and 225 amino acids (SHOXb). Both transcripts a and b share a common 5'end, but have a different last 3'exon, a finding suggestive of the use of alternative splicing signals. A complete alignment between the two cDNAs and the sequenced genomic DNA from cosmids LL0YNCO3''M''15D10 and LL0YNC3''M''34F5 was achieved, allowing establishment of the exon-intron structure (FIG. 4). The gene is composed of 6 exons ranging in size from 58 bp (exon III) to 1146 bp (exon Va). Exon I contains a CpG-island, the start codon and the 5' region. A stop codon as well as the 3'-noncoding region is located in each of the alternatively spliced exons Va and Vb.

EXAMPLE 3

[0083]Two cDNAs have been identified which map to the 160 kb region identified as critical for short stature. These cDNAs correspond to the genes SHOX and pET92. The cDNAs were identified by the hybridization of subclones of the cosmids to cDNA libraries.

[0084]Employing the set of cosmid clones with complete coverage of the critical region has now provided the genetic material to identify the causative gene. Positional cloning projects aimed at the isolation of the genes from this region are done by exon trapping and cDNA selection techniques. By virtue of their location within the pseudoautosomal region, these genes can be assumed to escape X-inactivation and to exert a dosage effect.

[0085]The cloning of the gene leading to short stature when absent (haploid) or deficient, represents a further step forward in diagnostic accuracy, providing the basis for mutational analysis within the gene by e.g. single strand conformation polymorphism (SSCP). In addition, cloning of this gene and its subsequent biochemical characterization has opened the way to a deeper understanding of biological processes involved in growth control.

[0086]The DNA sequences of the present invention provide a first molecular test to identify individuals with a specific genetic disorder within the complex heterogeneous group of patients with idiopathic short stature.

EXAMPLE 4

Expression Pattern of SHOXa and SHOXb

[0087]Northern blot analysis using single exons as hybridisation probes reveiled a different expression profile for every exon, strongly suggesting that the bands of different size and intensities represent cross-hybridisation products to other G,C rich gene sequences. To achieve a more realistic expression profile of both genes SHOXa and b, RT-PCR experiments on RNA from different tissues were carried out. Whereas expression of SHOXa was observed in skeletal muscle, placenta, pancreas, heart and bone marrow fibroblasts, expression of SHOXb was restricted to fetal kidney, skeletal muscle and bone marrow fibroblasts, with the far highest expression in bone marrow fibroblasts.

[0088]The expression of SHOXa in several cDNA libraries made of fetal brain, lung and muscle, of adult brain, lung and pituitary and of SHOXb in none of the tested libraries gives additional evidence that one spliced form (SHOXa) is more broadly expressed and the other (SHOXb) expressed in a predominantly tissue-specific manner.

[0089]To assess the transcriptional activity of SHOXa and SHOXb on the X and Y chromosome we used RT-PCR of RNA extracted from various cell lines containing the active X, the inactive X or the Y chromosome as the only human chromosomes. All cell lines revealed an amplification product of the expected length of 119 bp (SHOXa) and 541 bp (SHOXb), providing clear evidence that both SHOXa and b escape X-inactivation.

[0090]SHOXa and SHOXb encode novel homeodomain proteins. SHOX is highly conserved across species from mammalian to fish and flies. The very 5' end and the very 3' end--besides the homeodomain--are likely conserved regions between man and mouse, indicating a functional significance. Differences in those amino acid regions have not been allowed to accumulate during evolution between man and mouse.

Experimental Procedures:

a) 5' and 3'RACE

[0091]To clone the 5' end of the SHOXa and b transcripts, 5'RACE was performed using the constructed `Marathon cDNA libraries`. The following oligonucleotide primers were used: SHOX B rev, GAAAGGCATCCGTAAGGCTCCC (position 697-718, reverse strand [r]) and the adaptor primer AP1. PCR was carried out using touchdown parameters: 94° C. for 2 min, 94° C. for 30 sec, 70° C. for 30 sec, 72° C. for 2 min for 5 cycles. 94° C. for 30 sec, 66° C. for 30 sec, 72° C. for 2 min for 5 cycles. 94° C. for 30 sec, 62° C. for 30 sec, 72° C. for 2 min for 25 cycles. A second round of amplification was performed using 1/100 of the PCR product and the following nested oligonucleotide primers: SHOX A rev, GACGCCTTTATGCATCTGATTCTC (position 617-640 r) and the adaptor primer AP2. PCR was carried out for 35 cycles with an annealing temperature of 60° C.

[0092]To clone the 3' end of the SHOXa and b transcripts, 3'RACE was performed as previously described (Frohman et al., 1988) using oligo(dT)adaptor primed first strand cDNA. The following oligonucleotide primers were used: SHOX A for, GAATCAGATGCATAAAGGCGTC (position 619-640) and the oligo(dT)adaptor. PCR was carried out using following parameters: 94° C. for 2 min, 94° C. for 30 sec, 62° C. for 30 sec, 72° C. for 2 min for 35 cycles. A second round of amplification was performed using 1/100 of the PCR product and the following nested oligonucleotide primers: SHOX B for, GGGAGCCTTACGGATGCCTTTC (position 697-718) and the oligo(dT)adaptor. PCR was carried out for 35 cycles with annealing temperature of 62° C.

[0093]To validate the sequences of SHOXa and SHOXb transcripts, PCR was performed with a 5' oligonucleotide primer and a 3' oligonucleotide primer. For SHOXa the following primers were used: G310 for, AGCCCCGGCTGCTCGCCAGC (position 59-78) and SHOX D rev, CTGCGCGGCGGGTCAGAGCCCCAG (position 959-982 r). For SHOXb the following primers were used: G310 for, AGCCCCGGCTGCTCGCCAGC and SHOX2A rev, GCCTCAGCAGCAAAGCAAGATCCC (position 1215-1238 r). Both PCRs were carried out using touchdown parameters: 94° C. for 2 min, 94° C. for 30 sec, 70° C. for 30 sec, 72° C. for 2 min for 5 cycles. 94° C. for 30 sec, 68° C. for 30 sec, 72° C. for 2 min for 5 cycles. 94° C. for 30 sec, 65° C. for 30 sec, 72° C. for 2 min for 35 cycles. Products were gel-purified and cloned for sequencing analysis.

b) SSCP Analysis

[0094]SSCP analysis was performed on genomic amplified DNA from patients according to a previously described method (Orita et al., 1989). One to five μl of the PCR products were mixed with 5 μl of denaturation solution containing 95% Formamid and 10 mM EDTA pH8 and denaturated at 95° C. for 10 min. Samples were immediately chilled on ice and loaded on a 10% Polyacryamidgel (Acrylamide:Bisacryamide=37.5:1 and 29:1; Multislotgel, TGGE base, Qiagen) containing 2% glycerol and 1×TBE. Gels were run at 15° C. with 500V for 3 to 5 hours and silver stained as described in TGGE handbook (Qiagen, 1993).

c) Cloning and Sequencing of PCR Products

[0095]PCR products were cloned into pMOSBlue using the pMOSBlueT- Vector Kit from Amersham. Overnight cultures of single colonies were lysed in 100 μl H₂O by boiling for 10 min. The lysates were used as templates for PCRs with specific primers for the cloned PCR product. SSCP of PCR products allowed the identification of clones containing different alleles. The clones were sequenced with CY5 labelled vector primers Uni and T7 by the cycle sequencing method described by the manufacturer (ThermoSequenase Kit (Amersham)) on an ALF express automated sequencer (Pharmacia).

d) PCR Screening of cDNA Libraries

[0096]To detect expression of SHOXa and b, a PCR screening of several cDNA libraries and first strand cDNAs was carried out with SHOXa and b specific primers. For the cDNA libraries a DNA equivalent of 5×10⁸ pfu was used. For SHOXa, primers SHOX E rev, GCTGAGCCTGGACCTGTTGGAAAGG (position 713-737 r) and SHOX a for were used. For SHOXb, the following primers were used: SHOX B for and SHOX2A rev. Both PCRs were carried out using touchdown parameters: 94° C. for 2 min; 94° C. for 30 sec, 68° C. for 30 sec, 72° C. for 40 sec for 5 cycles. 94° C. for 30 sec, 65° C. for 30 sec, 72° C. for 40 sec for 5 cycles. 94° C. for 30 sec, 62° C. for 30 sec, 72° C. for 40 sec for 35 cycles.

e) PCR Screening of cDNA Libraries

[0097]To detect expression of SHOXa and b, a PCR screening of several cDNA libraries and first strand cDNAs was carried out with SHOXa and b specific primers. For the cDNA libraries a DNA equivalent of 5×10⁸ pfu was used. For SHOXa, primers SHOX E rev, GCTGAGCCTGGACCTGTTGGAAAGG (position 713-737 r) and SHOX a for were used. For SHOXb, the following primers were used: SHOX B for and SHOX2A rev. Both PCRs were carried out using touchdown parameters: 94° C. for 2 min; 94° C. for 30 sec, 68° C. for 30 sec, 72° C. for 40 sec for 5 cycles. 94° C. for 30 sec, 65° C. for 30 sec, 72° C. for 40 sec for 5 cycles. 94° C. for 30 sec, 62° C. for 30 sec, 72° C. for 40 sec for 35 cycles.

EXAMPLE 5

Expression Pattern of OG12, the Putative Mouse Homolog of Both SHOX and SHOT

[0098]In situ hybridisation on mouse embryos ranging from day 5 p.c. and day 18.5 p.c., as well as on fetal and newborn animals was carried out to establish the expression pattern. Expression was seen in the developing limb buds, in the mesoderm of nasal processes which contribute to the formation of the nose and palate, in the eyelid, in the aorta, in the developing female gonads, in the developing spinal cord (restricted to differentiating motor neurons) and brain. Based on this expression pattern and on the mapping position of its human homolog SHOT, SHOT represents a likely candidate for the Cornelia de Lange syndrome which includes short stature.

EXAMPLE 6

Isolation of a Novel SHOX-Like Homeobox Gene on Chromosome Three, SHOT, Being Related to Human Growth/Short Stature

[0099]A new gene called SHOT (for SHOX-homolog on chromosome three) was isolated in human, sharing the most homology with the murine OG12 gene and the human SHOX gene. The human SHOT gene and the murine OG12 genes are highly homologous, with 99% identity at the protein level. Although not yet proven, due to the striking homology between SHOT and SHOX ( identity within the homeodomain only), it is likely that SHOT is also a gene likely involved in short stature or human growth.

[0100]SHOT was isolated using primers from two new human ESTs (HS 1224703 and HS 126759) from the EMBL database, to amplify a reverse-transcribed RNA from a bone marrow fibroblast line (Rao et al, 1997). The 5' and 3' ends of SHOT were generated by RACE-PCR from a bone marrow fibroblast library that was constructed according to Rao et al., 1997. SHOT was mapped by FISH analysis to chromosome 3q25/q26 and the murine homolog to the syntenic region on mouse chromosome 3. Based on the expression pattern of OG12, its mouse homolog, SHOT represents a candidate for the Cornelia Lange syndrome (which shows short stature and other features, including craniofacial abnormalities) mapped to this chromosomal interval on 3q25/26.

EXAMPLE 7

Searching for Mutations in Patients with Idiopathic Short Stature

[0101]The DNA sequences of the present invention are used in PCR, LCR, and other known technologies to determine if such individuals with short stature have small deletions or point mutations in the short stature gene.

[0102]A total of initially 91 (in total 250 individuals) unrelated male and female patients with idiopathic short stature (idiopathic short stature has an estimated incidence of 2-2.5% in the general population) were tested for small rearrangements or point mutations in the SHOXa gene. Six sets of PCR primers were designed not only to amplify single exons but also sequences flanking the exon and a small part of the 5'UTR. For the largest exon, exon one, two additional internal-exon primers were generated. Primers used for PCR are shown in table 2.

[0103]Single strand conformation polymorphism (SSCP) of all amplified exons ranging from 120 to 295 bp in size was carried out. Band mobility shifts were identified in only 2 individuals with short stature (Y91 and A1). Fragments that gave altered SSCP patterns (unique SSCP conformers) were cloned and sequenced. To avoid PCR and sequencing artifacts, sequencing was performed on two strands using two independent PCR reactions. The mutation in patient Y91 resides 28bp 5' of the start codon in the 5'UTR and involves a cytidine-to-guanine substitution. To find out if this mutation represents a rare polymorphism or is responsible for the phenotype by regulating gene expression e.g. though a weaker binding of translation initiation factors, his parents and a sister were tested. As both the sister and father with normal height also show the same SSCP variant (data not shown), this base substitution represents a rare polymorphism unrelated to the phenotype.

[0104]Cloning and sequencing of a unique SSCP conformer for patient A1 revealed a cytidine-to-thymidine base transition (nucleotide 674) which introduces a termination codon at amino-acid position 195 of the predicted 225 and 292 amino-acid sequences, respectively. To determine whether this nonsense mutation is genetically associated with the short stature in the family, pedigree analysis was carried out. It was found that all six short individuals (defined as height below 2 standard deviations) showed an aberrant SSCP shift and the cytidine-to-thymidine transition. Neither the father, nor one aunt and maternal grandfather with normal height showed this mutation, indicating that the grandmother has transferred the mutated allele onto two of her daughters and her two grandchildren. Thus, there is concordance between the presence of the mutant allele and the short stature phenotype in this family.

[0105]The identical situation as indicated above was found in another short stature patient of Japanese origin.

EXAMPLE 8

[0106]The DNA sequences of the present invention are used to characterize the function of the gene or genes. The DNA sequences can be used as search queries for data base searching of nucleic acid or amino acid databases to identify related genes or gene products. The partial amino acid sequence of SHOX93 has been used as a search query of amino acid databases. The search showed very high homology to many known homeobox proteins. The cDNA sequences of the present invention can be used to recombinantly produce the peptide. Various expression systems known to those skilled in the art can be used for recombinant protein production.

[0107]By conventional peptide synthesis (protein synthesis according to the Merrifield method), a peptide having the sequence CSKSFDQKSKDGNGG (SEQ ID NO: 42) was synthesized and polyclonal antibodies were derived in both rabbits and chicken according to standard protocols.

REFERENCES

[0108]The following references are herein incorporated by reference.

[0109]Ashworth A, Rastan S, Lovell-Badge R, Kay G (1991): X-chromosome inactivation may explain the difference in viability of X0 humans and mice. Nature 351: 406-408.

[0110]Ballabio A, Bardoni A, Carrozzo R, Andria G, Bick D, Campbell L, Hamel B, Ferguson-Smith MA, Gimelli G, Fraccaro M, Maraschio P, Zuffardi O, Guilo S, Camerino G (1989): Contiguous gene syndromes due to deletions in the distal short arm of the human X chromosome. Proc Natl Acad Sci USA 86:10001-10005.

[0111]Blagowidow N, Page D C, Huff D, Mennuti M T (1989): Ullrich-Tumer syndrome in an XY female fetus with deletion of the sex-determining portion of the Y chromosome. Am. J. med. Genet. 34:159-162.

[0112]Cantrell M A, Bicknell J N, Pagon R A et al. (1989): Molecular analysis of 46,XY females and regional assignment of a new Y-chromosome-specific probe. Hum. Genet. 83: 88-92.

[0113]Connor J M, Loughlin S A R (1989): Molecular genetics of Turner's syndrome. Acta Pediatr. Scand. (Suppl.) 356: 77-80.

[0114]Disteche C M, Casanova M, Saal H, Friedmen C, Sybert V, Graham J, Thuline H, Page D C, Fellous M (1986): Small deletions of the short arm of the Y-chromosome in 46,XY females. Proc Natl Acad Sci USA 83:7841-7844.

[0115]Ferguson-Smith M A (1965): Karyotype-phenotype correlations in gonadal dysgenesis and their bearing on the pathogenesis of malformations. J. med. Genet. 2: 142-155.

[0116]Ferrari D, Kosher R A, Dealy C N (1994): Limb mesenchymal cells inhibited from undergoing cartilage differentiation by a tumor promoting phorbol ester maintain expression of the homeobox-containing gene MSX1 and fail to exhibit gap junctional communication. Biochemical and Biophysical Research Communications. 205(1): 429-434.

[0117]Fischer M, Bur-Romero P, Brown L G et al. (1990): Homologous ribosomal protein genes in the human X- and Y-chromosomes escape from X-inactivation and possible implementation for Turner syndrome. Cell 63: 1205-1218.

[0118]Freund C, Horsford D J, McInnes R R (1996): Transcription factor genes and the developing eye: a genetic perspective. Hum Mol Genet 5: 1471-1488.

[0119]Gehring W J, Qian Y Q, Billeter M, Furukubo-Tokunaga K, Schier A F, Resendez-Perez D, Affolter M, Otting G, Wuthrich K (1994): Homeodomain-DNA recognition. Cell 78: 211-223.

[0120]Gough N M, Gearing D P, Nicola N A, Baker E, Pritchard M, Callen D F, Sutherland G R (1990). Localization of the human GM-CSF receptor gene to the X-Y pseudoautosomal region. Nature 345: 734736

[0121]Grumbach M M, Conte F A (1992): Disorders of sexual differentiation. In: Williams textbook of endocrinology, 8th edn., edited by Wilson J D, Foster D W, pp. 853-952, Philadelphia, W B Saunders.

[0122]Hall J G, Gilchrist D M (1990): Turner syndrome and its variants. Pedriatr. Clin. North Am. 37: 1421-1436.

[0123]Henke A, Wapenaar M, van Ommen G-J, Maraschio P, Camerino O, Rappold G A (1991): Deletions within the pseudoautosomal region help map three new markers and indicate a possible role of this region in linear growth. Am J Hum Genet 49:811-819.

[0124]Hernandez D, Fisher E M C (1996): Down syndrome genetics: unravelling a multifactorial disorder. Hum Mol Genet 5:1411-1416.

[0125]Kenyon C (1994): If birds can fly, why can't we? Homeotic genes and evolution. Cell 78: 175-180.

[0126]Krumlauf R (1994): Hox genes in vertebrate development. Cell 78: 191-201.

[0127]Kulharya A S, Roop H, Kukolich M K, Nachtman R G, Belmont J W, Garcia-Heras J (1995): Mild phenotypic effects of a de novo deletion Xpter→Xp22.3 and duplication 3pter→3p23. Am J Med Genet 56:16-21.

[0128]Lawrence P A, Morata G (1994): Homeobox genes: their function in Drosophila segmentation and pattern formation. Cell 78: 181-189.

[0129]Lehrach H, Drmnac R, Hoheisel J D, Larin Z, Lemon G, Monaco A P, Nizetic D, et a,. Hybridization finger printing in genome mapping and sequencing. In Davies K E, Tilghman S, Eds. Genome Analysis 1990: 39-81 Cold Spring Harbor, N.Y.

[0130]Levilliers J, Quack B, Weissenbach J, Petit C (1989): Exchange of terminal portions of X- and Y-chromosomal short arms in human XY females. Proc Natl Acad Sci USA 86:2296-2300.

[0131]Lichter P, Cremer T, Human Cytogenetics: A practical Approach, IRL Press 1992, Oxford, New York, Tokyo

[0132]Lippe B M (1991): Turner Syndrome. Endocrinol Metab Clin North Am 20: 121-152. Magenis R E, Tochen M L Holahan K P, Carey T, Allen L, Brown M G (1984): Turner syndrome resulting from partial deletion of Y-chromosome short arm: localization of male determinants. J Pediatr 105: 916-919.

[0133]Nelson D L, Ballabio A, Cremers F, Monaco A P, Schlessinger D (1995).--Report of the sixth international workshop on the X chromosome mapping. Cytogenet. Cell Genet. 71: 308-342

[0134]Ogata T, Goodfellow P, Petit C, Aya M, Matsuo N (1992): Short stature in a girl with a terminal Xp deletion distal to DXYS15: localization of a growth gene(s) in the pseudoautosomal region. J Med Genet 29:455-459.

[0135]Ogata T, Tyler-Smith C, Purvis-Smith S, Turner G (1993): Chromosomal localisation of a gene(s) for Turner stigmata on Yp. J. Med. Genet. 30: 918-922.

[0136]Ogata T, Yoshizawa. A, Muroya K, Matsuo N, Fukushima Y, Rappold GA, Yokoya S (1995): Short stature in a girl with partial monosomy of the pseudoautosomal region distal to DXYS15: further evidence for the assignment of the critical region for a pseudoautosomal growth gene(s). J Med Genet 32:831-834.

[0137]Ogata T, Matsuo N (1995): Turner syndrome and female sex chromosome aberrations: deduction of the principle factors involved in the development of clinical features. Hum. Genet. 95: 607-629.

[0138]Orita M, Suzuki Y, Sekiya T and Hayashi K (1989): Rapid and sensitive detection of point mutations and polymorphisms using the polymerase chain reaction. Genomics 5:874-879.

[0139]Pohlschmidt M, Rappold G A, Krause M, Ahlert D, Hosenfeld D, Weissenbach J, Gal A (1991): Ring Y chromosome: Molecular characterization by DNA probes. Cytogenet Cell Genet 56:65-68.

[0140]Qiagen (1993) TGGE Handbook, Diagen GmbH, TGMA 4112 3/93.

[0141]Rao E, Weiss B, Mertz A et al. (1995): Construction of a cosmid contig spanning the short stature candidate region in the pseudoautosomal region PAR 1. in: Turner syndrome in a life span perspective: Research and clinical aspects. Proceedings of the 4th International Symposium on Turner Syndrome, Gothenburg, Sweden, 18-21 May, 1995., edited by Albertsson-Wikland K, Ranke M B, pp. 19-24, Elsevier.

[0142]Rao E, Weiss B, Fukami M, Rump A, Niesler B, Mertz A, Muroya K, Binder G, Kirsch S, Winkelmann M, Nordsiek G, Heinrich U, Breuning M H, Ranke M B, Rosenthal A, Ogata T, Rappold G A (1997): Pseudoautosomal deletions encompassing a novel homeobox gene cause growth failure in idiopathic short stature and Turner syndrome. Nature Genet 15:54-62

[0143]Rappold G A (1993): The pseudoautosomal region of the human sex chromosomes. Hum Genet 92:315-324.

[0144]Rappold G A, Willson T A, Henke A, Gough N M (1992): Arrangement and localization of the human GM-CSF receptor α chain gene CSF2RA within the X-Y pseudoautosomal region. Genomics 14:455-461.

[0145]Ried K, Mertz A, Nagaraja R, Trusnich M, Riley J, Anand R, Page D, Lehrach H., Elliso J, Rappold G A (1995): Characterization of a yeast artificial chromosome contig spanning the pseudoautosomal region. Genomics 29:787-792.

[0146]Robinson A (1990): Demography and prevalence of Turner syndrome. In: Turner Syndrome., edited by Rosenfeld R G, Grumbach M M, pp. 93-100, New York, Marcel Dekker.

[0147]Rosenfeld R G (1992): Turner syndrome: a guide for physicians. Second edition. The Turner's Syndrome Society.

[0148]Rosenfeld R G, Tesch L-G, Rodriguez-Rigau L J, McCauley E, Albertsson-Wikland K, Asch R, Cara J, Conte F, Hall J G, Lippe B, Nagel T C, Neely E K, Page D C, Ranke M, Saenger P, Watkins J M, Wilson D M (1994): Recommendations for diagnosis,treatment, and management of individuals with Turner syndrome. The Endocrinologist 4(5): 351-358.

[0149]Rovescalli A C, Asoh S, Nirenberg M (1996): Cloning and characterization of four murine homeobox genes. Proc Natl Acad Sci USA 93:10691-10696.

[0150]Schaefer L, Ferrero G B, Grillo A, Bassi M T, Roth E J, Wapenaar M C, van Ommen G-J B, Mohandas T K, Rocchi M, Zoghbi H Y, Ballabio A (1993): A high resolution deletion map of human chromosome Xp22. Nature genetics 4: 272-279.

[0151]Shalet S M (1993): Leukemia in children treated with growth hormone. Journal of Pediatric Endocrinology 6: 109-11

[0152]Vimpani G V, Vimpani A F, Lidgard G P, Cameron E H D, Farquhar J W (1977) Prevalence of severe growth hormone deficiency. Br Med J. 2: 427-430

[0153]Zinn A R, Page D C, Fisher E M C (1993): Turner syndrome: the case of the missing sex chromosome. TIG 9 (3): 90-93.

Sequence CWU 1

55160PRTHomo sapiens 1Gln Arg Arg Ser Arg Thr Asn Phe Thr Leu Glu Gln Leu Asn Glu Leu 1 5 10 15Glu Arg Leu Phe Asp Glu Thr His Tyr Pro Asp Ala Phe Met Arg Glu 20 25 30Glu Leu Ser Gln Arg Leu Gly Leu Ser Glu Ala Arg Val Gln Val Trp 35 40 45Phe Gln Asn Arg Arg Ala Lys Cys Arg Lys Gln Glu 50 55 602209DNAHomo sapiens 2ggatttatga atgcaaagag aagcgcgagg acgtgaagtc ggaggacgag gacgggcaga 60ccaagctgaa acagaggcgc agccgcacca acttcacgct ggagcagctg aacgagctcg 120agcgactctt cgacgagacc cattaccccg acgccttcat gcgcgaggag ctcagccagc 180gcctggggct ctccgaggcg cgcgtgcag 2093368DNAHomo sapiens 3gtgatccacc cgcgcgcacg ggccgtcctc tccgcgcggg gagacgcgcg catccaccag 60ccccggctgc tcgccagccc cggccccagc catggaagag ctcacggctt ttgtatccaa 120gtcttttgac cagaaaagca aggacggtaa cggcggaggc ggaggcggcg gaggtaagaa 180ggattccatt acgtaccggg aagttttgga gagcggactg gcgcgctccc gggagctggg 240gacgtcggat tccagcctcc aggacatcac ggagggcggc ggccactgcc cggtgcattt 300gttcaaggac cacgtagaca atgacaagga gaaactgaaa gaattcggca ccgcgagagt 360ggcagaag 368458DNAHomo sapiens 4gtttggttcc agaaccggag agccaagtgc cgcaaacaag agaatcagat gcataaag 58589DNAHomo sapiens 5gcgtcatctt gggcacagcc aaccacctag acgcctgccg agtggcaccc tacgtcaaca 60tgggagcctt acggatgcct ttccaacag 8961166DNAHomo sapiens 6gtccaggctc agctgcagct ggaaggcgtg gcccacgcgc acccgcacct gcacccgcac 60ctggcggcgc acgcgcccta cctgatgttc cccccgccgc ccttcgggct gcccatcgcg 120tcgctggccg agtccgcctc ggccgccgcc gtggtcgccg ccgccgccaa aagcaacagc 180aagaattcca gcatcgccga cctgcggctc aaggcgcgga agcacgcgga ggccctgggg 240ctctgacccg ccgcgcagcc ccccgcgcgc ccggactccc gggctccgcg caccccgcct 300gcaccgcgcg tcctgcactc aaccccgcct ggagctcctt ccgcggccac cgtgctccgg 360gcaccccggg agctcctgca agaggcctga ggagggaggc tcccgggacc gtccacgcac 420gacccagcca gaccctcgcg gagatggtgc agaaggcgga gcgggtgagc ggccgtgcgt 480ccagcccggg cctctccaag gctgcccgtg cgtcctggga ccctggagaa gggtaaaccc 540ccgcctggct gcgtcttcct ctgctatacc ctatgcatgc ggttaactac acacgtttgg 600aagatcctta gagtctattg aaactgcaaa gatcccggag ctggtctccg atgaaaatgc 660catttcttcg ttgccaacga ttttctttac taccatgctc cttccttcat cccgagaggc 720tgcggaacgg gtgtggattt gaatgtggac ttcggaatcc caggaggcag gggccgggct 780ctcctccacc gctcccccgg agcctcccag gcagcaataa ggaaatagtt ctctggctga 840ggctgaggac gtgaaccgcg ggctttggaa agggagggga gggagacccg aacctcccac 900gttgggactc ccacgttccg gggacctgaa tgaggaccga ctttataact tttccagtgt 960ttgattccca aattgggtct ggttttgttt tggattggta tttttttttt tttttttttt 1020tgctgtgtta caggattcag acgcaaaaga cttgcataag agacggacgc gtggttgcaa 1080ggtgtcatac tgatatgcag cattaacttt actgacatgg agtgaagtgc aatattataa 1140atattataga ttaaaaaaaa aatagc 11667625DNAHomo sapiens 7atggagtttt gctcttgtcg cccaggctgg agtataatgg catgatctcg actcactgca 60acctccgcct cccgagttca agcgattctc ctgcctcagc ctcccgagta gctgggatta 120caggtgccca ccaccatgtc aagataatgt ttgtattttc agtagagatg gggtttgacc 180atgttggcca ggctggtctc gaactcctga cctcaggtga tccacccgcc ttagcctccc 240aaagtgctgg gatgacaggc gtgagcccct gcgcccggcc tttgtaactt tatttttaat 300tttttttttt ttttaagaaa gacagagtct tgctctgtca cccaggctgg agcacactgg 360tgcgatcata gctcactgca gcctcaaact cctgggctca agcaatcctc ccacctcagc 420ctcctgagta gctgggacta caggcaccca ccaccacacc cagctaattt ttttgatttt 480tactagagac gggatcttgc tttgctgctg aggctggtct tgagctcctg agctccaaag 540atcctctcac ctccacctcc caaagtgtta gaattacaag catgaaccac tgcccgtggt 600ctccaaaaaa aggactgtta cgtgg 625815577DNAHomo sapiensmisc_feature(3844)..(4068)pET92 region (first part) 8ctctccctgt tgtgtctctc tttctctctc tccatctctc tccgtctttc cccctctgtc 60tctttctctg tctccatccc tctgtctctc cctttctctc tgtctttcct tgtctctctc 120tttctctctc tctctccatc tctctctctc ccggtctctc tctctccatc tccccgtctc 180tccgtttctc tctctgcctc tccctgtctg tctctctctt tgtgtgtgtt acacacaccc 240caacccaccg tcactcatgt ccccccactg ctgtgccatc tcacacaagt tcacagctca 300gctgtcatcc tgggtcccca ggccccgccg gggaggaaga tgcgccgtgg ggttacggga 360ggaaggggac tccgggcctc ctggtgcccc actttatttg cagaaggtcc ttggcaggaa 420ccgtgacgcg tttggtttcc aggacttgga aaacgaattt caggtcgcga tggcgagcac 480cggcttcccc tgaagcacat tcaatagcga gaggcgggag ggagcgagca ggagcatccc 540accatgaaaa ccaaaaacac aagtattttt ttcacccggt aaatacccca gacgccaggg 600tgacagcgcg gcgctaaggg aggaggcctc gcgccggggt ccgccgggat ctggcgcggg 660cggaaagaat atagatcttt acgaaccgga tctcccgggg acctgggctt ctttctgcgg 720gcgctggaaa cccgggaggc ggccccgggg atcctcggcc tccgccgccg ccgcctccca 780agcgcccgcg tcccggtttg gggacacccg gccccttctt ctcactttcg gggattctcc 840agccgcgttc catctcacca actctccatc caagggcgcg ccgccaccaa cttggagctc 900atcttctccc aaaatcgtgc gtccccgggg cgcccgggtc ccccccctcg ccatctcaac 960cccggcgcga cccgggcgct tcctggaaag atccaggcgc cgggctctgc gctcctcccg 1020ggagcgaggg cggccggaca actgggaccc tcctctctcc agccgtgaac tccttgtctc 1080tctgtctctc tctgcaggaa aactggagtt tgcttttcct ccggccacgg aaagaacgcg 1140ggtaacctgt gtggggggct cgggcgcctg cgcccccctc ctgcgcgcgc gctctccctt 1200ccaaaaatgg gatctttccc ccttcgcacc aaggtgtacg gacgccaaac agtgatgaaa 1260tgagaagaaa gccaattgcc ggcctggggg gtgggggaga cacagcgtct ctgcgtgcgt 1320ccgccgcgga gcccggagac cagtaattgc accagacagg cagcgcatgg ggggctgggc 1380gaggtcgccg cgtataaata gtgagatttc caatggaaag gcgtaaataa cagcgctggt 1440gatccacccg cgcgcacggg ccgtcctctc cgcgcgggga gacgcgcgca tccaccagcc 1500ccggctgctc gccagccccg gccccagcca tggaagagct cacggctttt gtatccaagt 1560cttttgacca gaaaagcaag gacggtaacg gcggaggcgg aggcggcgga ggtaagaagg 1620attccattac gtaccgggaa gttttggaga gcggactggc gcgctcccgg gagctgggga 1680cgtcggattc cagcctccag gacatcacgg agggcggcgg ccactgcccg gtgcatttgt 1740tcaaggacca cgtagacaat gacaaggaga aactgaaaga attcggcacc gcgagagtgg 1800cagaaggtaa gttcctttgc gcgccggctc caggggggcc ctcctggggt tcggcgcctc 1860ctcgccacgg agtcggcccc gcgcgcccct cgctgtgcac atttgcagct cccgtctcgc 1920cagggtaagg cccgggccgt caggctttgc ctaagaaagg aaggaaggca ggagtggacc 1980cgaccggaga cgcgggtggt gggtagcggg gtgcgggggg acccagggag ggtcgcagcg 2040ggggccgcgc gcgtgggcac cgacacggga aggtcccggg ctggggtgga tccgggtggc 2100tgtgcctgaa gccgtagggc ctgagatgtc tttttcattt tctttttctt tcctttcctt 2160tttttgtttg tttgtttgtt tgtttgagac agagtctcgc tctgtccccc aggctggagt 2220gcagtggtgc gatctcggct cactgcaacc tccgcctcct gggttcaagc gattctcctg 2280cctcagcctc cccagtagct gggattacag gcatgcacca ccacgcctgg ctaatttttg 2340tgcttttagt aaagacgggg attcaccatg ttggccaggc tggtctcgaa ctcctgacct 2400caggtgatcc acccgcctcg gcctcccaaa gtgctgggat gacaggcgtg aggcaccgcg 2460cccggcctgg gtcctgacgg cttaggatgt gtgtttctgt ctctgcctgt ctgccttgta 2520tttacggtca cccagacgca cagaggagcc gtctccacgc gccttcccag cgctcagcgc 2580ctgccgggcc cccggagatc acgggaagac tcgaggctgc gtggtaggag acgggaaggc 2640cccgggtcag ctcggttctg tttcncttta aggaaccctt cattattatt tcattgtttt 2700cctttgaacg tcgaggcttg atcttggcga aagctgttgg gtccataaaa accactcccg 2760tgagcggagg tggccgggat ctggatgggg cgcgaggggc cccggggaag ctggcggctt 2820cgcgggcgcg tcctaagtca aggttgtcag agcgcagccg gttgtgcgcg gcccgggggn 2880agctcccctc tggcccttcc tcctgagacc tcagtggtgg gtcgtcccgt ggtggaaatc 2940ggggagtaag aggctcagag agaggggctg gccccgggga tctctgtgca cacacgacaa 3000ctgggcggca tacatcttaa gaataaaatg ggctggctgt gtcggggcac agctggagac 3060ggctatggac gcctgttatg ttttcattac aaagacgcag agaatctagc ctcggctttt 3120gctgattcgc aaagttgagg tgcgagggtg aatgccccaa aggtaattct tcctaagact 3180ctggggctac ctgctctccg gggccctgca tttggggtgt ggagtggccc cgggaaatag 3240cccttgtatt cgtaggaggc accaggcagc ttcccaaggc cctgactttg tcgaagcaga 3300aagctgtggc tacggtttac aaagcagtcc ccggtttctg accgtctaag aggcaggagc 3360ccagcctgcc tttgacagtg agaggagttc ctccctacac actgctgcgg gcacccggca 3420ctgtaattca tacacagaga gttggccttc ctggacgcaa ggctgggagc cgcttgaggg 3480cctgcgtgta atttaagagg gttcgcangc gcccggcggc cgcttctgnt ggggttgctt 3540tttggttgtc cttcngcaaa caccgttttg ctcctctngn aactctctct tnctcccccn 3600tggccngtng gacccgggna ngagcaaagt gtcctccaga ccnttttgaa angtgagagg 3660aaaataaaga ccaggccaaa nngacccagg gccacaggag aggagacaga gagtccccgt 3720tacattttnc cccttggctg ggtgcagaaa gacccccggg ccaggactgc cacccaggct 3780actatttatt catcagatcc aagttaaatc gaggttggag ggcaggggag agtctgaggt 3840taccgtggaa gcctggagtt tttgggnaac agcgtgtccc cgccgagcct gggagcccgt 3900gggttctgca aagcctgcgg gtgtttgagg actttgaaga ccagtttgtc agttgggctc 3960aattncctgg ggttcagact tagagaaatg aaggagggag agctggggtc gtctccagga 4020aacgattcac ttggggggaa ggaatggagt gttcttgcag gcacatgtct gttaggaggt 4080gaaacagaat gtgaaatcca cgttggagta agcgtccagc gctgaatgta gctcggggtg 4140gggtgggagg gccctggtgt ggatcgtgga aggnaagaaa gacagaacag ggtgctagta 4200tttaccccgt tnccctgtag acaccctgga tttgtcagct ttgcaagctt cttggttgca 4260gcggccttgc ctgtgcccct ttgagactgt ttccagacta aacttccaaa tgtcagcccc 4320ttacccttga cagcaaggga catctcatta gggcatcgcg tgcttctcat ctgtgnctca 4380gcaggcccng agataggaan cangaggggc ngttggnaga tgcncacttc caccagccct 4440gggnttgaag gggangcgan gggangacna ccttttanct taaacccctn gagcttggtn 4500cagagaggnc tgaatgtcta aaatgaggaa gaaaaggttt ttcacctgga aacgcttgag 4560ggctgagtct tctgcccntt ctgacntccc ccagcaaata cagacaggtc accaanctac 4620tggagatgag aaagtgccat ttttggcaca ctctggtggg gtaggtgccc gaccgcgtgt 4680gaaaaangtg ggaannggag agatttctgn cgcacgcggt tcagccccca ggcgcggntg 4740gcngcattcn aggntactca gacgcggttc tgctgttctg ctgagaaaca ggcttcgggt 4800aggggctcct agctccgcca gatcgcggag ggacccccag ccctcctgcg ctgcagcggt 4860ggggatagcg tctctccgta ggcctagaat ctgcaacccg ccccgggtcc tccccgtgtc 4920cttcccgggc gtcccgccgg ggatcccaca gttggcagct cttcctcaaa ttctttccct 4980taaaaatagg atttgacacc ccactctcct taaaaaaaaa aaataagaaa aaaaggttag 5040gttatgtcaa cagaggtgaa gtggataatt gaggaaacga ttctgagatg aggccaagaa 5100aacaacgctc gtgcaaagcc caggtttttg ggaaagcagc gagtatcctc ctcggctttt 5160gcgttatgga ccccacgcag tttttgcgtc aaagcgcatt ggttttcgag ggcccccttt 5220ccaccgcggg atgcacgaag gggttcgcca cgttgcgcaa aacctccccg gcctcagccc 5280tgtgccctcc gctccccacg cagggattta tgaatgcaaa gagaagcgcg aggacgtgaa 5340gtcggaggac gaggacgggc agaccaagct gaaacagagg cgcagccgca ccaacttcac 5400gctggagcag ctgaacgagc tcgagcgact ttttgacgag acccattacc ccgacgcctt 5460catgcgcgag gagctcagcc agcgcctggg gctttccgag gcgcgcgtgc aggtaggaac 5520ccgggggcgg gggcgggggg cccggagcca tcgcctggtc ctcgggagcg cacagcacgc 5580gtacagccac ctgcgcccgg gccgccgccg tccccttccc ggagcgcggg gaggttgggt 5640gagggacggg ctggggttcc tggacttttg gagacgcctg aggcctgtag gatgggttca 5700ttgcgtttgt ttttcaccaa cagcaaacaa atatatatac atatatatta tacaaataac 5760aaataaatat atatgttata cagatgggta tattgtatat attatagata tttgttcgtc 5820cttggtgcaa agacacccgg tgaacccata tattggctcc tgactgcctt cggttcccct 5880gggattggtt ataggggcaa cacatgcaaa caaaactttc cctggattat acttaggaga 5940cgaagctaca gatgcgtttg atccagagtg ttttacaaga tttttcattt aaaaaaaaat 6000gtgtcttttg gcccctgatt cccctccgtc ttcccgtgtg gctgcattga aaaggtttcc 6060ttaggatgaa aggagagggg tgtcctctgt ccctaggtgg agagaaacag ggtcttctct 6120ttcctccgtt ttttcaccta ccgtttctat ctccctcctc ccctctccag ccctgtcctc 6180tgctacaaac caccccctcc tccctccggc tgtggggagc gcaggagcac gttgggcatc 6240tggatgagcg gnagactatt agcggggcac gggggctccc cgaggagcgc gcgaattcac 6300gctgccccat gagaccaggc accggggggc ggaggggcct tgggtgtccg cagagggacg 6360ggcgggcaga gccttcctcc gcattctaaa cattcactta aaggtatgag tttantttca 6420ggggtgctgc tgggagagcc tccaaatggc ttcttccagc ccctgcctga cagttcagct 6480cccctggaag gtcaactcct ctagtccttt ctcctggttc tgggcaggac agaagtgggg 6540ggagggagag agagagagag agagagagag acggtcagga tccccggacc ctggggaacc 6600cgtcaaaaat aaatgaaatt aagattgccg accagagaga gaaccgtgac aaagcaaacg 6660gcgttcaaag caaagagacg aactgaaagc ccgttcccgt aggactggtt atgaggtcaa 6720cacattcaaa cacagcttgc tctggatttt gctgagcaga ggaagataca gatgcatttg 6780atccaaagtg tgttacatct ttcattatat gtgtgtctat atatataaac atatataaat 6840atataaacat acataaatgt atgtaaatat atataatcta tatacatata taaatatata 6900aacacatata taatatataa atctataaac atatataata tataaacata aatatataaa 6960catatataat atataaatat attaacatat ataaaatatg tataaatata tataaacata 7020taaacatata taaatatata aacatataaa tatataaaca tatataaata tatacaaaca 7080tattgtatat atataaatat atataaaaac atatatatac atataaaaat atatataaac 7140atatatacat ataaagaaat atatataaac atatatacat ataaatatac atatataaac 7200atatatatac ataaaatata tataaacata tatacatata aaaatatata tatattaaca 7260tatatataca tataaaaata tatatattaa catatatata catataaaaa tatatatata 7320tttttggccc ctgattccct tcggttcctg tgggatgggt gattgagtca acacattcaa 7380acacaacttt tccatcgatg ttgcttagga gatgaggata cagatgcgtt tgatggagag 7440ggttttacaa gctctttcat ttaaatatat atatatatat atatatattt tttggctcct 7500gattctcttc cgtcttccca tgtggctgca ttttaaaagg cttccctaag atcgttacga 7560ttaaatcaac cctccccagg catctttacc gagggctgtg gtccccaaag cgatacagcc 7620caggagggag agaggctttg gtgacttgga ggaaggactg tgtccctcct tagggcgtct 7680gtggcctcag tgagggaagg aagctgcatc agacaggggt ttcctcgctg tccacccctc 7740tggcagaaga tggattgggc tgccccgnta taaattaatg aaaagattaa agtttcgcta 7800aaggggacat cgagtttatg tgtcatctcc tggtgntctg tgtgccntgg gatnctgcaa 7860tatatcccan ngcccttgat gnnntactgt ttnctataaa aanntaaatn tacttgtnna 7920atttaanttc cnnnacacta tttnctttcc nngtnagtct nattanccga ncgagagcan 7980cgnttagttn cagctngcgg aaaattggtt gtggggtgtg tgcggacccc ngagnaacgc 8040ccnntaaaat naaagacaaa ntcnggggac aagnctnggg ggttatcgnn attgcnnagg 8100ggtcgncatg aaaantttaa cgacggtaaa taataataaa aanncaaaca tgggaatgnc 8160aataaaagac ataattctcc nnatcgccgc ggggggaaag gatcctatag taaaggcgag 8220tgcgctttga ggggtcataa aaatcaatta gttccaacac ccacgtcccg cgttgagggg 8280acggggacga gcagggacag aaaaagaaac catatttgaa tcccatctct ctgtgaattc 8340ttgggtcaca tgcgtctcag tacagcccgt cccgtgctgt gaccggatag agtttcaatt 8400tactgtggaa atttgctgta aataaattga gcatccgata gaagctgttg ctgattaacc 8460ttttattttt agcgtggccc tgcaaagtcg tatcacccag ctgtcaggct tctaatcgaa 8520agttatgaga ccacggtgag gggcaggcgg taatttaatt acaacaaata tctttgggtt 8580tatggcgcag agctaaatta aatgtcatta ttcactgtct gtnaatggna aatcaaaann 8640ggaaatcgca nttacggnca tttgggnnaa angaaagcgg ggnagtgctc tttaatngaa 8700nngaaataac tgtcttaagc agtgtcacac acttcactta ccatattcgn ggcctnaatt 8760ggaanntgga tcgtnngaat cactccnaag actngattta ttangcgctt cacgncagcn 8820nggcntaatt catcnacttn ngtattcttc atcnnnnatt tttttttttc ctctcnngcc 8880gtgttnngaa gggagagtga atgaggcttt ccacgtttca ggaggatttt cttttttgaa 8940aaatgccctt ccagaggctt ttgggtggct ggcttgcttt ctgggccctg gaggangaca 9000ggcggangag tccaggtggg catggagagg cacagtggca ggtcacctgg atggtcagtg 9060gaggtggagg tctgaaggcg ccagctttgg aaattattgg tgaatttcga tgtcagcacc 9120aggncagggg cctttttggc gggggtgtga ggganggatg anctttgctg ggaaanncag 9180gatcaggttc tccaggcgca ctgcagcccg gtaggaccca ctttggaaat gaaaagccag 9240ttnccgaaag ctgggctgga agcttccgtg ttgggttcaa gagcaagttc acgttgcgct 9300gtgtagactc ctggctgctc ccaaactctg agggttttct gaggttccct tcataggggc 9360accggccctg ggccatgcac agtgcgtaag ggtggctgtg ggccgaggga cccagcacgt 9420gttttgccca caacagccgg agtgactggt tcactcaccg ccttggcgga ggacgcctgt 9480tctctggacg aatcatttct cttgggtggt gactgccttg tgggtcaagg tgcaggtttt 9540ctgccacaga aaacctgtta ggaggaatta agcgactaag actgtcaggg aggtggtggt 9600gggggangag gnagggggtg gtgtccagat taccaggcat aggctaaact gcctgcactc 9660tccagctggt ctgtctgtgg aggaggggat tgtcaatact gggagagcag aggaggctcg 9720taggaggtga gagggggtgg aatttgcatg caaatcttca catgaggcct gtgtgaattt 9780ctccagcctc ctgagggtcc cctgcgctat tgcactcaac ttcttgatag tttaccccaa 9840gactcagaag tccttagagg ggcagaatgc ccccaccaca aagcctgcta tccttgggcg 9900tcctcaggac ccttggtcat gaatgggacc ctttcatgta tggggaccct tggtaatatg 9960aatgggacgc cttcagctcc ccagggcttc cgaggaggcc gagaagggca aagacacttc 10020cgaggaggcc gagaagggca aagacatttt ctgggcttgg tgtgtcaaga gctagattgg 10080agaaggggct ggatttggaa ctctttagcc atcagctcac cctctccgtt tgtggctaaa 10140gtctgaaggt ggaaacttcg gttctcctac agggtctaca ggagttgggg ggcggggcgc 10200ccacacagaa cgctggaaag ttcgacagtc cacttccact ggctcggaac tcactttttc 10260accttaagtt catcagcggt aacgcatagg tctcacttag gcagggcacg gatgatttaa 10320caatttctac ttctaggtca ggtgcggtgg ctcacacctc taatcccagc actttgggag 10380gcccaggagg gtggatcgct tgaggtcagg agtttgagac cagcctggcc aacatggtga 10440aaccccgtct ctactaaaat acgaaaatta gccaggcatg gtggtgagca cctgtaattc 10500cagctactcg ggaggctgag gcaggagaat cgcttgaacc tgggaggtgg acgttgcagt 10560gaggtgagat cacaccactg cactccagcc tggatgagag agcaagactc tgtctcaaaa 10620acaaaataaa acaaaaacaa aacaaaaatc aaaaaagaaa acccaatttc cagttctagg 10680ccaggtgcag tggctcacgc ctgtcatccc agcactttgg gaggcccagg agggtggatc 10740gcttgaggtc aggagttcga gaccagcctg gccaacatgg tgaaacccca tctttactaa 10800aaatacaaac gttagctggg tgtggtggtg tgcgcctgta atcccagcta ctcgggaagc 10860tgaggctgga gaattgcttg aatctgggag gtggaggttg cagggaggcg agatagtgcc 10920actgcagtcc agcctggacc agagagcaag actccgtctc aaaaacaaaa gaaagcaaaa 10980acaaaaaaca agagaccagc ctggccaaca tggtgaaacc gcgtctttac taaaatacaa 11040aattagccgg gcatggtggt gggcacctgt agtcccagct actcgggagg ctgaggcagg 11100agaatggctt gaacctggga ggtggagctt gcagtgagcc gagatagtgc cactgcactc 11160cagcctgggc gacagagcga gacttgattt cagaaccacc accaccacaa caaaacaaaa 11220caaaaaatcc aaaaaaaccc caatttccag tactaggtag tcagtgatgc agggctggag 11280acagaggggc ggtaagtgtc tgggcgccca ccatcagtca cctcccagct cccangaggt 11340gcaaagtgct tggttcagcc tcatgggaag gatgctccct ggggaggctg ggctgggttc 11400acagggctct tcacatctct ctctgcttct nccccaaggt ttggttncca gaaccggaga 11460gccaagtgcc gncaaacaag agaatcagat gcataaaggt gggtgtcggg actgggggga 11520cctgaagctg ggggatcctg ctccaggagg gatggggtcg acaaggtgct ggctacaccc 11580aggaccacca cactgacacc tgctcccttt ggacacaggc gtcatcttgg gcacagccaa 11640ccacctagac gcctgccnga gtggcaccct acgtcaacat gggagcctta cggatgcctt 11700tccaacaggt agctcacttt ttcttcctct gnaagatccc tagggacctg ctgctccctt 11760cccctttccc

ctatttgctg ccgcatcctg acactcctag tccctccctg cccctgcaga 11820cttctcagct ggcccttaga aaaaaagcct cttttccgag gaggcattta caggcacctt 11880ggcacctatg aaatcaggct gggccaggcg gggtggctca cacctgtcat cccagcactt 11940tgggaggcca aggttaggag tttgagacca gcctggacaa catagcaaaa gcctgtctct 12000actaaaaata caaaaaaaaa ttaacaggga gtggtggtgg gcacctgtaa tcccagctac 12060ttgggaggct gaggcaggag aatcacttga acccgggagg ccgaggttgc ggtgagccga 12120gatcgtgcca ttgcactcca ggctgggcga cagagtgaga ctctgtctca aaaaataaat 12180aaataaataa atgtaaaaaa ataaaaatag gtcgggcacg gtggctcacg tctgtaatcc 12240cagcactttg gaaggccgag gtgggtggat gacagggtca agagattgag accatcctgg 12300ccaacatggc aaaatgccgt ctctactaaa aaatacaaaa attaggcggg cgtggtggcg 12360ggtgcctgta atcccagcta ctcgggaggc tgaggcagga gaatcggttg aacccgggat 12420gcggaggttg cagtgagcgg agatcacatc actgcactcc aggctgggca acaagagcga 12480aactgcgtct tacaataaat aaatagataa ataaataaac aaataaactt tactttagaa 12540acaaatccct gtccgtgttt gtcttttcac ctgtcctgca gggaaaacaa aacataaaat 12600gtcaaggcaa atagtagtga tttcattccg ggaaaaagaa agtggatgtt tgccttcacc 12660ctttctcgtc cttcctctgg tgctcctcan ggcccanggg nagagggtgg aaagtncaga 12720ggaagaaaga cggggctggg ggggggggtc cgtggggacc caggcaggca tgttcccnat 12780ttccntgtct tcacnttcaa agnaggggcc cctcgnctct ggaatgaggc ctacggtttc 12840ctttcccnga agagttnccc ctttgtgagc ttacggcttc ggagtgaacc tcggtgcaac 12900ctgttattaa aacacacaga ggctaatgcc agcaaaaaca cgccccccgc tcctggtttc 12960agagggaaga aaaaaattca taagcacggc catgcttttc taataaaaat tcattaaata 13020atcgttataa gggatgaagc cgggagggga gaggagagga acacaatcaa gagactttct 13080ttgaactttt tctccctgct tcaaatacaa agcaatcttc tgtgggcctg ggcctggggg 13140gtttccccct ttctctgcag cccattggga ggaagaaaat gcttccctga angttgctgc 13200aaaattgttt ctgtttttct tttctttttc tttttttttt ttttttgaga cggagtctcg 13260ctctgtcacc aggctggagt gcaatggtat gatctcagct cactgcaacc tccacgttcc 13320tgtttcaagt cattctcctg cctcagcctc ctgagtagct gggactacag gcgcccgcca 13380ccacgcccgg ctagtgtttg tatttttaga aaagacaggg tttccccatg ttggccaggc 13440tggtcttgaa ctcctgtcct caagtgatct gcctgcctcg gcctcccaaa gtgctgtgtt 13500tctgtttttc tttccccgct ttcttaggag gccatcggga agaataaaat gctttccttg 13560aagttgatgc aaaattgttt ctgtttttct tttctctttt ctttcttttt gagatggagt 13620ctcgctcttt cacccaggct ggagggcagt ggcgcgacct cggctcactg caacctccgc 13680ctcccgggtt caagcgattc tcctgcctca gcctccggag tagctgggat tacaggcacc 13740tgccactatg cctggctaat tttattattt ttagtagaga cggggtttca ccatgttggc 13800caggctggtc tcaaactcct gacctcaggt gatccgcccg cctcgcctcc caaagtgatg 13860ggatgancag gncatngagc ncaccgtgcc cggccctcta actctttacc agacataaag 13920tctccnnttc ccctttctaa atgtatatat tgtgttttta aaagttaaca gcagggatcc 13980cacctcattn ccccgctnct ctccccaaga cctgtcctgc acgttgcaca cagcaggtgt 14040gccctggaca tatcccaaac ccacgctgaa agaaagaggg tctcactaca cgtatgatat 14100ctgtgnatcc tttaaacatc tccgtggctt ccaggcaaca cagccataaa taggaatctc 14160atgtctgaca tgataccggg accatgtatg ggnaaattct gggtgtgaag ttccagctac 14220ccccgcagag gcanccattg cataccctcc agaaactccc ctgccgttnc aagccaaaga 14280cacaacacaa acagcntccg agagagggtg tcattgaaaa tcaataccat cataagagca 14340cacagcaccg tctttctctt ctgcccgttg atacacaatt atgagcaatt tgctaacact 14400gacaactcgt ggcaagaaca ggtcgtgttg atacggttgc ctcgtgagga cccatctgtc 14460ttctggggtc ttgcctggaa cggagatcgg agttcagggt ggctaataga atcattactc 14520acctagggac acagaatnat gagggttacc cccagttaag tgcatacagt caaacggacg 14580gctgctctgg aaggtacagt gacgtgaaca gcttttatga aatgcctaga tctggacctt 14640ccatacctga gccaccgttc caaagcactg ggcgtttttc agatactttc atgagaaatg 14700ttgtcaacac cgcaagtttg cagtacacag tctgaaagat attcttgtat atgtagatgt 14760ctgtagatgc cctgaaggtg tgtagacttt agacacccag aaggtgtgta gatgtctgta 14820gacaccttct atgtgtgtag atgtctgtag acgccctgca ggtgtgtaga tatatctaga 14880tggtctgcct gtgtatgata caggctaaaa agacatttgt ggtggacact agttgattat 14940ttaggactat gagatgggaa aggaagnagc aaccagcagt gaaaggcatg tggtgggtgg 15000ggggttggca ttgcagtggg gtcctcntga ngcaggtgac acccactata gggctgccct 15060tggnatggac gctttgtnga agctgtttga tttcaccaca ccaagcctgg aggcacggac 15120attccaggat ggtgaggagt ctgcaaagga ggagattgga ggaggtgcaa tatccctaga 15180gtacgagaga tgagatagga gagctgtata aatagcacta ccagccggat gcggtggctc 15240acgcctgtca tcccagcact ttaggaggct gaggcaggcg gatcacctga ggtcaggagt 15300tccagaacag cctggccaac acaatgaaac cccatcttta ctaaaaatac aagattagct 15360gggcacggtg tctcacgcct gtcatccctg cactttggga ggtcgaggtg cgcagatcat 15420gaggtcagtt tggccaacgc ggcgaaaccc cgtctctact aaaaatacaa aaaagtagcc 15480gggcgtggtg gtgggcacct gtagtcccag ctactaggga ggctgaggca ggagaatcgc 15540ttgaacccgg atgcggacat tgcagtgagc cgagatc 155779753DNAHomo sapiens 9cgtggaagcc tggagttttt gggaacagcg tgtccccgcc gagcctggga gcccgtgggt 60tctgcaaagc ctgcgggtgt ttgaggactt tgaagaccag tttgtcagtt gggctcaatt 120cctggggttc agacttagag aaatgaagga gggagagctg gggtcgtctc caggaaacga 180ttcacttggg gggaaggaat ggagtgttct tgcaggcaca tgtctgttag gaggtgaaac 240agaatgtgaa atccacgttg gagtaagcgt ccagcgctga atgtagctcg gggtggggtg 300ggagggccct ggtgtggatc gtggaaggaa gaaagacaga acagggtgct agtatttacc 360ccgttccctg tagacaccct ggatttgtca gctttgcaag cttcttggtt gcagcggcct 420tgcctgtgcc cctttgagac tgtttccaga ctaaacttcc aaatgtcagc cccttaccct 480tgacagcaag ggacatctca ttagggcatc gcgtgcttct catctgtgct cagcaggccc 540gagataggaa cagaggggcg ttggagatgc cacttccacc agccctgggt tgaaggggag 600cgagggagac accttttact taaacccctg agcttggtca gagaggctga atgtctaaaa 660tgaggaagaa aaggtttttc acctggaaac gcttgagggc tgagtcttct gcccttctga 720ctcccccagc aaatacagac aggtcaccaa cta 753101895DNAHomo sapiensCDS(91)..(966) 10gtgatccacc cgccgcacgg gccgtcctct ccgcgcgggg agacgcgcgc atccaccagc 60cccggctgct cgccagcccc ggccccagcc atg gaa gag ctc acg gct ttt gta 114 Met Glu Glu Leu Thr Ala Phe Val 1 5tcc aag tct ttt gac cag aaa agc aag gac ggt aac ggc gga ggc gga 162Ser Lys Ser Phe Asp Gln Lys Ser Lys Asp Gly Asn Gly Gly Gly Gly 10 15 20ggc ggc gga ggt aag aag gat tcc att acg tac cgg gaa gtt ttg gag 210Gly Gly Gly Gly Lys Lys Asp Ser Ile Thr Tyr Arg Glu Val Leu Glu 25 30 35 40agc gga ctg gcg cgc tcc cgg gag ctg ggg acg tcg gat tcc agc ctc 258Ser Gly Leu Ala Arg Ser Arg Glu Leu Gly Thr Ser Asp Ser Ser Leu 45 50 55cag gac atc acg gag ggc ggc ggc cac tgc ccg gtg cat ttg ttc aag 306Gln Asp Ile Thr Glu Gly Gly Gly His Cys Pro Val His Leu Phe Lys 60 65 70gac cac gta gac aat gac aag gag aaa ctg aaa gaa ttc ggc acc gcg 354Asp His Val Asp Asn Asp Lys Glu Lys Leu Lys Glu Phe Gly Thr Ala 75 80 85aga gtg gca gaa ggg att tat gaa tgc aaa gag aag cgc gag gac gtg 402Arg Val Ala Glu Gly Ile Tyr Glu Cys Lys Glu Lys Arg Glu Asp Val 90 95 100aag tcg gag gac gag gac ggg cag acc aag ctg aaa cag agg cgc agc 450Lys Ser Glu Asp Glu Asp Gly Gln Thr Lys Leu Lys Gln Arg Arg Ser105 110 115 120cgc acc aac ttc acg ctg gag cag ctg aac gag ctc gag cga ctc ttc 498Arg Thr Asn Phe Thr Leu Glu Gln Leu Asn Glu Leu Glu Arg Leu Phe 125 130 135gac gag acc cat tac ccc gac gcc ttc atg cgc gag gag ctc agc cag 546Asp Glu Thr His Tyr Pro Asp Ala Phe Met Arg Glu Glu Leu Ser Gln 140 145 150cgc ctg ggg ctc tcc gag gcg cgc gtg cag gtt tgg ttc cag aac cgg 594Arg Leu Gly Leu Ser Glu Ala Arg Val Gln Val Trp Phe Gln Asn Arg 155 160 165aga gcc aag tgc cgc aaa caa gag aat cag atg cat aaa ggc gtc atc 642Arg Ala Lys Cys Arg Lys Gln Glu Asn Gln Met His Lys Gly Val Ile 170 175 180ttg ggc aca gcc aac cac cta gac gcc tgc cga gtg gca ccc tac gtc 690Leu Gly Thr Ala Asn His Leu Asp Ala Cys Arg Val Ala Pro Tyr Val185 190 195 200aac atg gga gcc tta cgg atg cct ttc caa cag gtc cag gct cag ctg 738Asn Met Gly Ala Leu Arg Met Pro Phe Gln Gln Val Gln Ala Gln Leu 205 210 215cag ctg gaa ggc gtg gcc cac gcg cac ccg cac ctg cac ccg cac ctg 786Gln Leu Glu Gly Val Ala His Ala His Pro His Leu His Pro His Leu 220 225 230gcg gcg cac gcg ccc tac ctg atg ttc ccc ccg ccg ccc ttc ggg ctg 834Ala Ala His Ala Pro Tyr Leu Met Phe Pro Pro Pro Pro Phe Gly Leu 235 240 245ccc atc gcg tcg ctg gcc gag tcc gcc tcg gcc gcc gcc gtg gtc gcc 882Pro Ile Ala Ser Leu Ala Glu Ser Ala Ser Ala Ala Ala Val Val Ala 250 255 260gcc gcc gcc aaa agc aac agc aag aat tcc agc atc gcc gac ctg cgg 930Ala Ala Ala Lys Ser Asn Ser Lys Asn Ser Ser Ile Ala Asp Leu Arg265 270 275 280ctc aag gcg cgg aag cac gcg gag gcc ctg ggg ctc tgacccgccg 976Leu Lys Ala Arg Lys His Ala Glu Ala Leu Gly Leu 285 290cgcagccccc cgcgcgcccg gactcccggg ctccgcgcac cccgcctgca ccgcgcgtcc 1036tgcactcaac cccgcctgga gctccttccg cggccaccgt gctccgggca ccccgggagc 1096tcctgcaaga ggcctgagga gggaggctcc cgggaccgtc cacgcacgac ccagccagac 1156cctcgcggag atggtgcaga aggcggagcg ggtgagcggc cgtgcgtcca gcccgggcct 1216ctccaaggct gcccgtgcgt cctgggaccc tggagaaggg taaacccccg cctggctgcg 1276tcttcctctg ctatacccta tgcatgcggt taactacaca cgtttggaag atccttagag 1336tctattgaaa ctgcaaagat cccggagctg gtctccgatg aaaatgccat ttcttcgttg 1396ccaacgattt tctttactac catgctcctt ccttcatccc gagaggctgc ggaacgggtg 1456tggatttgaa tgtggacttc ggaatcccag gaggcagggg ccgggctctc ctccaccgct 1516cccccggagc ctcccaggca gcaataagga aatagttctc tggctgaggc tgaggacgtg 1576aaccgcgggc tttggaaagg gaggggaggg agacccgaac ctcccacgtt gggactccca 1636cgttccgggg acctgaatga ggaccgactt tataactttt ccagtgtttg attcccaaat 1696tgggtctggt tttgttttgg attggtattt tttttttttt ttttttttgc tgtgttacag 1756gattcagacg caaaagactt gcataagaga cggacgcgtg gttgcaaggt gtcatactga 1816tatgcagcat taactttact gacatggagt gaagtgcaat attataaata ttatagatta 1876aaaaaaaaat agcaaaaaa 189511292PRTHomo sapiens 11Met Glu Glu Leu Thr Ala Phe Val Ser Lys Ser Phe Asp Gln Lys Ser 1 5 10 15Lys Asp Gly Asn Gly Gly Gly Gly Gly Gly Gly Gly Lys Lys Asp Ser 20 25 30Ile Thr Tyr Arg Glu Val Leu Glu Ser Gly Leu Ala Arg Ser Arg Glu 35 40 45Leu Gly Thr Ser Asp Ser Ser Leu Gln Asp Ile Thr Glu Gly Gly Gly 50 55 60His Cys Pro Val His Leu Phe Lys Asp His Val Asp Asn Asp Lys Glu 65 70 75 80Lys Leu Lys Glu Phe Gly Thr Ala Arg Val Ala Glu Gly Ile Tyr Glu 85 90 95Cys Lys Glu Lys Arg Glu Asp Val Lys Ser Glu Asp Glu Asp Gly Gln 100 105 110Thr Lys Leu Lys Gln Arg Arg Ser Arg Thr Asn Phe Thr Leu Glu Gln 115 120 125Leu Asn Glu Leu Glu Arg Leu Phe Asp Glu Thr His Tyr Pro Asp Ala 130 135 140Phe Met Arg Glu Glu Leu Ser Gln Arg Leu Gly Leu Ser Glu Ala Arg145 150 155 160Val Gln Val Trp Phe Gln Asn Arg Arg Ala Lys Cys Arg Lys Gln Glu 165 170 175Asn Gln Met His Lys Gly Val Ile Leu Gly Thr Ala Asn His Leu Asp 180 185 190Ala Cys Arg Val Ala Pro Tyr Val Asn Met Gly Ala Leu Arg Met Pro 195 200 205Phe Gln Gln Val Gln Ala Gln Leu Gln Leu Glu Gly Val Ala His Ala 210 215 220His Pro His Leu His Pro His Leu Ala Ala His Ala Pro Tyr Leu Met225 230 235 240Phe Pro Pro Pro Pro Phe Gly Leu Pro Ile Ala Ser Leu Ala Glu Ser 245 250 255Ala Ser Ala Ala Ala Val Val Ala Ala Ala Ala Lys Ser Asn Ser Lys 260 265 270Asn Ser Ser Ile Ala Asp Leu Arg Leu Lys Ala Arg Lys His Ala Glu 275 280 285Ala Leu Gly Leu 290121354DNAHomo sapiensCDS(91)..(765) 12gtgatccacc cgccgcacgg gccgtcctct ccgcgcgggg agacgcgcgc atccaccagc 60cccggctgct cgccagcccc ggccccagcc atg gaa gag ctc acg gct ttt gta 114 Met Glu Glu Leu Thr Ala Phe Val 1 5tcc aag tct ttt gac cag aaa agc aag gac ggt aac ggc gga ggc gga 162Ser Lys Ser Phe Asp Gln Lys Ser Lys Asp Gly Asn Gly Gly Gly Gly 10 15 20ggc ggc gga ggt aag aag gat tcc att acg tac cgg gaa gtt ttg gag 210Gly Gly Gly Gly Lys Lys Asp Ser Ile Thr Tyr Arg Glu Val Leu Glu 25 30 35 40agc gga ctg gcg cgc tcc cgg gag ctg ggg acg tcg gat tcc agc ctc 258Ser Gly Leu Ala Arg Ser Arg Glu Leu Gly Thr Ser Asp Ser Ser Leu 45 50 55cag gac atc acg gag ggc ggc ggc cac tgc ccg gtg cat ttg ttc aag 306Gln Asp Ile Thr Glu Gly Gly Gly His Cys Pro Val His Leu Phe Lys 60 65 70gac cac gta gac aat gac aag gag aaa ctg aaa gaa ttc ggc acc gcg 354Asp His Val Asp Asn Asp Lys Glu Lys Leu Lys Glu Phe Gly Thr Ala 75 80 85aga gtg gca gaa ggg att tat gaa tgc aaa gag aag cgc gag gac gtg 402Arg Val Ala Glu Gly Ile Tyr Glu Cys Lys Glu Lys Arg Glu Asp Val 90 95 100aag tcg gag gac gag gac ggg cag acc aag ctg aaa cag agg cgc agc 450Lys Ser Glu Asp Glu Asp Gly Gln Thr Lys Leu Lys Gln Arg Arg Ser105 110 115 120cgc acc aac ttc acg ctg gag cag ctg aac gag ctc gag cga ctc ttc 498Arg Thr Asn Phe Thr Leu Glu Gln Leu Asn Glu Leu Glu Arg Leu Phe 125 130 135gac gag acc cat tac ccc gac gcc ttc atg cgc gag gag ctc agc cag 546Asp Glu Thr His Tyr Pro Asp Ala Phe Met Arg Glu Glu Leu Ser Gln 140 145 150cgc ctg ggg ctc tcc gag gcg cgc gtg cag gtt tgg ttc cag aac cgg 594Arg Leu Gly Leu Ser Glu Ala Arg Val Gln Val Trp Phe Gln Asn Arg 155 160 165aga gcc aag tgc cgc aaa caa gag aat cag atg cat aaa ggc gtc atc 642Arg Ala Lys Cys Arg Lys Gln Glu Asn Gln Met His Lys Gly Val Ile 170 175 180ttg ggc aca gcc aac cac cta gac gcc tgc cga gtg gca ccc tac gtc 690Leu Gly Thr Ala Asn His Leu Asp Ala Cys Arg Val Ala Pro Tyr Val185 190 195 200aac atg gga gcc tta cgg atg cct ttc caa cag atg gag ttt tgc tct 738Asn Met Gly Ala Leu Arg Met Pro Phe Gln Gln Met Glu Phe Cys Ser 205 210 215tgt cgc cca ggc tgg agt ata atg gca tgatctcgac tcactgcaac 785Cys Arg Pro Gly Trp Ser Ile Met Ala 220 225ctccgcctcc cgagttcaag cgattctcct gcctcagcct cccgagtagc tgggattaca 845ggtgcccacc accatgtcaa gataatgttt gtattttcag tagagatggg gtttgaccat 905gttggccagg ctggtctcga actcctgacc tcaggtgatc cacccgcctt agcctcccaa 965agtgctggga tgacaggcgt gagcccctgc gcccggcctt tgtaacttta tttttaattt 1025tttttttttt ttaagaaaga cagagtcttg ctctgtcacc caggctggag cacactggtg 1085cgatcatagc tcactgcagc ctcaaactcc tgggctcaag caatcctccc acctcagcct 1145cctgagtagc tgggactaca ggcacccacc accacaccca gctaattttt ttgattttta 1205ctagagacgg gatcttgctt tgctgctgag gctggtcttg agctcctgag ctccaaagat 1265cctctcacct ccacctccca aagtgttaga attacaagca tgaaccactg cccgtggtct 1325ccaaaaaaag gactgttacg tggaaaaaa 135413225PRTHomo sapiens 13Met Glu Glu Leu Thr Ala Phe Val Ser Lys Ser Phe Asp Gln Lys Ser 1 5 10 15Lys Asp Gly Asn Gly Gly Gly Gly Gly Gly Gly Gly Lys Lys Asp Ser 20 25 30Ile Thr Tyr Arg Glu Val Leu Glu Ser Gly Leu Ala Arg Ser Arg Glu 35 40 45Leu Gly Thr Ser Asp Ser Ser Leu Gln Asp Ile Thr Glu Gly Gly Gly 50 55 60His Cys Pro Val His Leu Phe Lys Asp His Val Asp Asn Asp Lys Glu 65 70 75 80Lys Leu Lys Glu Phe Gly Thr Ala Arg Val Ala Glu Gly Ile Tyr Glu 85 90 95Cys Lys Glu Lys Arg Glu Asp Val Lys Ser Glu Asp Glu Asp Gly Gln 100 105 110Thr Lys Leu Lys Gln Arg Arg Ser Arg Thr Asn Phe Thr Leu Glu Gln 115 120 125Leu Asn Glu Leu Glu Arg Leu Phe Asp Glu Thr His Tyr Pro Asp Ala 130 135 140Phe Met Arg Glu Glu Leu Ser Gln Arg Leu Gly Leu Ser Glu Ala Arg145 150 155 160Val Gln Val Trp Phe Gln Asn Arg Arg Ala Lys Cys Arg Lys Gln Glu 165 170 175Asn Gln Met His Lys Gly Val Ile Leu Gly Thr Ala Asn His Leu Asp 180 185 190Ala Cys Arg Val Ala Pro Tyr Val Asn Met Gly Ala Leu Arg Met Pro 195 200 205Phe Gln Gln Met Glu Phe Cys Ser Cys Arg Pro Gly Trp Ser Ile Met 210 215 220Ala2251432367DNAHomo sapiens 14tttctctgtc tccatccctc tgtctctccc tttctctctg tctttccttg tctctctctt 60tctctctctc tctccatctc tctctctccc tgtctctctc tctccatctc cccgtctctc 120cgtttctctc tctgcctctc cctgtctgtc tctctctttc tgtgtcttac acacacccca 180acccaccgtc actcatgtcc ccccactgct

gtgccatctc acacaagttc acagctcagc 240tgtcatcctg ggtccccagg ccccgccggg gaggaagatg cgccgtgggg ttacgggagg 300aaggggactc cgggcctcct ggtgccccac tttatttgca gaaggtcctt ggcaggaacc 360gtgacgcgtt tggtttccag gacttggaaa acgaatttca ggtcgcgatg gcgagcaccg 420gcttcccctg aagcacattc aatagcgaga ggcgggaggg agcgagcagg agcatcccac 480catgaaaacc aaaaacacaa gtattttttt cacccggtaa ataccccaga cgccagggtg 540acagcgcggc gctaagggag gaggcctcgc gccggggtcc gccgggatct ggcgcgggcg 600gaaagaatat agatctttac gaaccggatc tcccggggac ctgggcttct ttctgcgggc 660gctggagacc cgggaggcgg ccccggggat cctcggcctc cgccgccgcc gcctcccaag 720cgcccgcgtc ccggtttggg gacacccggc cccttcttct cactttcggg gattctccag 780ccgcgttcca tctcaccaac tctccatcca agggcgcgcc gccaccaact tggagctcat 840cttctcccaa gatcgtgcgt ccccggggcg cccgggtccc ccccctcgcc atctcaaccc 900cggcgcgacc cgggcgcttc ctggaaagat ccaggcgccg ggctctgcgc tcctcccggg 960agcgagggcg gccggacgac tgggaccctc ctctctccag ccgtgaactc cttgtctctc 1020tgtctctctc tgcaggaaaa ctggagtttg cttttcctcc ggccacggag agaacgcggg 1080taacctgtgt ggggggctcg ggcgcctgcg cccccctcct gcgcgcgcgc tctcccttcc 1140aaaaatggga tctttccccc ttcgcaccaa ggtgtacgga cgccaaacag tgatgaaatg 1200agaagaaagc caattgccgg cctggggggt gggggagaca cagcgtctct gcgtgcgtcc 1260gccgcggagc ccggagacca gtaattgcac cagacaggca gcgcatgggg ggctgggcga 1320ggtcgccgcg tataaatagt gagatttcca atggaaaggc gtaaataaca gcgctggtga 1380tccacccgcg cgcacgggcc gtcctctccg cgcggggaga cgcgcgcatc caccagcccc 1440ggctgctcgc cagccccggc cccagccatg gaagagctca cggcttttgt atccaagtct 1500tttgaccaga aaagcaagga cggtaacggc ggaggcggag gcggcggagg taagaaggat 1560tccattacgt accgggaagt tttggagagc ggactggcgc gctcccggga gctggggacg 1620tcggattcca gcctccagga catcacggag ggcggcggcc actgcccggt gcatttgttc 1680aaggaccacg tagacaatga caaggagaaa ctgaaagaat tcggcaccgc gagagtggca 1740gaaggtaagt tcctttgcgc gccggctcca ggggggccct cctggggttc ggcgcctcct 1800cgccacggag tcggccccgc gcgcccctcg ctgtgcacat ttgcagctcc cgtctcgcca 1860gggtaaggcc cgggccgtca ggctttgcct aagaaaggaa ggaaggcagg agtggacccg 1920accggagacg cgggtggtgg gtagcggggt gcggggggac ccagggaggg tcgcagcggg 1980ggccgcgcgc gtgggcaccg acacgggaag gtcccgggct ggggtggatc cgggtggctg 2040tgcctgaagc cgtagggcct gagatgtctt tttcattttc tttttctttc ctttcctttt 2100tttgtttgtt tgtttgtttg tttgagacag agtctcgctc tgtcccccag gctggagtgc 2160agtggtgcga tctcggctca ctgcaacctc cgcctcctgg gttcaagcga ttctcctgcc 2220tcagcctccc cagtagctgg gattacaggc atgcaccacc acgcctggct aatttttgtg 2280cttttagtaa agacggggat tcaccatgtt ggccaggctg gtctcgaact cctgacctca 2340ggtgatccac ccgcctcggc ctcccaaagt gctgggatga caggcgtgag gcaccgcgcc 2400cggcctgggt cctgacggct taggatgtgt gtttctgtct ctgcctgtct gccttgtatt 2460tacggtcacc cagacgcaca gaggagccgt ctccacgcgc cttcccagcg ctcagcgcct 2520gccgggcccc cggagatcac gggaagactc gaggctgcgt ggtaggagac gggaaggccc 2580cgggtcagct cggttctgtt tcctttaagg aacccttcat tattatttca ttgttttcct 2640ttgaacgtcg aggcttgatc ttggcgaaag ctgttgggtc cataaaaacc actcccgtga 2700gcggaggtgg ccgggatctg gatggggcgc gaggggcccc ggggaagctg gcggcttcgc 2760gggcgcgtcc taagtcaagg ttgtcagagc gcagccggtt gtgcgcggcc cgggggagct 2820cccctctggc ccttcctcct gagacctcag tggtgggtcg tcccgtggtg gaaatcgggg 2880agtaagaggc tcagagagag gggctggccc cggggatctc tgtgcacaca cgacaactgg 2940gcggcataca tcttaagaat aaaatgggct ggctgtgtcg gggcacagct ggagacggct 3000atggacgcct gttatgtttt cattacaaag acgcagagaa tctagcctcg gcttttgctg 3060attcgcagag ttgaggtgcg agggtgaatg ccccaaaggt aattcttcct aagactctgg 3120ggctacctgc tctccggggc cctgcatttg gggtgtggag tggccccggg aaatagccct 3180tgtattcgta ggaggcacca ggcagcttcc caaggccctg actttgtcga agcagaaagc 3240tgtggctacg gtttacaaag cagtccccgg tttctgaccg tctaagaggc aggagcccag 3300cctgcctttg acagtgagag gagttcctcc ctacacactg ctgcgggcac ccggcactgt 3360aattcataca cagagagttg gccttcctgg acgcaaggct gggagccgct tgagggcctg 3420cgtgtaattt aagagggttc gcagcgcccg gcggccgctt ctgtggggtt gctttttggt 3480tgtccttcgc agacaccgtt ttgctcctct gaactctctc ttctccccct ggccgtggac 3540ccgggagagc aaagtgtcct ccagaccttt tgaaagtgag aggaaaataa agaccaggcc 3600aaagacccag ggccacagga gaggagacag agagtccccg ttacattttc cccttggctg 3660ggtgcagaaa gacccccggg ccaggactgc cacccaggct actatttatt catcagatcc 3720aagttaaatc gaggttggag ggcaggggag agtctgaggt taccgtggaa gcctggagtt 3780tttgggaaca gcgtgtcccc gccgagcctg ggagcccgtg ggttctgcaa agcctgcggg 3840tgtttgagga ctttgaagac cagtttgtca gttgggctca attcctgggg ttcagactta 3900gagaaatgaa ggagggagag ctggggtcgt ctccaggaaa cgattcactt ggggggaagg 3960aatggagtgt tcttgcaggc acatgtctgt taggaggtga aacagaatgt gaaatccacg 4020ttggagtaag cgtccagcgc tgaatgtagc tcggggtggg gtgggagggc cctggtgtgg 4080atcgtggaag gaagaaagac agaacagggt gctagtattt accccgttcc ctgtagacac 4140cctggatttg tcagctttgc aagcttcttg gttgcagcgg ccttgcctgt gcccctttga 4200gactgtttcc agactaaact tccaaatgtc agccccttac ccttgacagc aagggacatc 4260tcattagggc atcgcgtgct tctcatctgt gctcagcagg cccgagatag gaacagaggg 4320gcgttggaga tgccacttcc accagccctg ggttgaaggg gagcgaggga gacacctttt 4380acttaaaccc ctgagcttgg tcagagaggc tgaatgtcta aaatgaggaa gaaaaggttt 4440ttcacctgga aacgcttgag ggctgagtct tctgcccttc tgactccccc agcaaataca 4500gacaggtcac caactactgg agatgagaaa gtgccatttt tggcacactc tggtggggta 4560ggtgcccgac cgcgtgtgaa aaagtgggaa ggagagattt ctgcgcacgc ggttcagccc 4620ccaggcgcgg tggcgcattc aggtactcag acgcggttct gctgttctgc tgagaaacag 4680gcttcgggta ggggctccta gctccgccag atcgcggagg gacccccagc cctcctgcgc 4740tgcagcggtg gggatagcgt ctctccgtag gcctagaatc tgcaacccgc cccgggtcct 4800ccccgtgtcc ttcccgggcg tcccgccggg gatcccacag ttggcagctc ttcctcaaat 4860tctttccctt aaaaatagga tttgacaccc cactctcctt aaaaaaaaaa aataagaaaa 4920aaaggttagg ttatgtcaac agaggtgaag tggataattg aggaaacgat tctgagatga 4980ggccaagaaa acaacgctcg tgcaaagccc aggtttttgg gaaagcagcg agtatcctcc 5040tcggcttttg cgttatggac cccacgcagt ttttgcgtca aagcgcattg gttttcgagg 5100gccccctttc caccgcggga tgcacgaagg ggttcgccac gttgcgcaaa acctccccgg 5160cctcagccct gtgccctccg ctccccacgc agggatttat gaatgcaaag agaagcgcga 5220ggacgtgaag tcggaggacg aggacgggca gaccaagctg aaacagaggc gcagccgcac 5280caacttcacg ctggagcagc tgaacgagct cgagcgactc ttcgacgaga cccattaccc 5340cgacgccttc atgcgcgagg agctcagcca gcgcctgggg ctctccgagg cgcgcgtgca 5400ggtaggaacc cgggggcggg ggcggggggc ccggagccat cgcctggtcc tcgggagcgc 5460acagcacgcg tacagccacc tgcgcccggg ccgccgccgt ccccttcccg gagcgcgggg 5520aggttgggtg agggacgggc tggggttcct ggacttttgg agacgcctga ggcctgtagg 5580atgggttcat tgcgtttgtt tttcaccaac agcaaacaaa tatatataca tatatattat 5640acaaataaca aataaatata tatgttatac agatgggtat attgtatata ttatagatat 5700ttgttcgtcc ttggtgcaaa gacacccggt gaacccatat attggctcct gactgccttc 5760ggttcccctg ggattggtta taggggcaac acatgcaaac aaaactttcc ctggattata 5820cttaggagac gaagctacag atgcgtttga tccagagtgt tttacaagat ttttcattta 5880aaaaaaaatg tgtcttttgg cccctgattc ccctccgtct tcccgtgtgg ctgcattgaa 5940aaggtttcct taggatgaaa ggagaggggt gtcctctgtc cctaggtgga gagaaacagg 6000gtcttctctt tcctccgttt tttcacctac cgtttctatc tccctcctcc cctctccagc 6060cctgtcctct gctacaaacc accccctcct ccctccggct gtggggagcg caggagcacg 6120ttgggcatct ggatgagcgg agactattag cggggcacgg gggctccccg aggagcgcgc 6180gaattcacgc tgccccatga gaccaggcac cggggggcgg aggggccttg ggtgtccgca 6240gagggacggg cgggcagagc cttcctccgc attctaaaca ttcacttaaa ggtatgagtt 6300tatttcaggg gtgctgctgg gagagcctcc aaatggcttc ttccagcccc tgcctgacag 6360ttcagctccc ctggaaggtc aactcctcta gtcctttctc ctggttctgg gcaggacaga 6420agtgggggga gggagagaga gagagagaga gagagagacg gtcaggatcc ccggaccctg 6480gggaacccgt caaaaataaa tgaaattaag attgccgacc agagagagaa ccgtgacaaa 6540gcaaacggcg ttcaaagcaa agagacgaac tgaaagcccg ttcccgtagg actggttatg 6600aggtcaacac attcaaacac agcttgctct ggattttgct gagcagagga agatacagat 6660gcatttgatc caaagtgtgt tacatctttc attatatgtg tgtctatata tataaacata 6720tataaatata taaacataca taaatgtatg taaatatata taatctatat acatatataa 6780atatataaac acatatataa tatataaatc tataaacata tataatatat aaacataaat 6840atataaacat atataatata taaatatatt aacatatata aaatatgtat aaatatatat 6900aaacatataa acatatataa atatataaac atataaatat ataaacatat ataaatatat 6960acaaacatat tgtatatata taaatatata taaaaacata tatatacata taaaaatata 7020tataaacata tatacatata aagaaatata tataaacata tatacatata aatatacata 7080tataaacata tatatacata aaatatatat aaacatatat acatataaaa atatatatat 7140attaacatat atatacatat aaaaatatat atattaacat atatatacat ataaaaatat 7200atatatattt ttggcccctg attcccttcg gttcctgtgg gatgggtgat tgagtcaaca 7260cattcaaaca caacttttcc atcgatgttg cttaggagat gaggatacag atgcgtttga 7320tggagagggt tttacaagct ctttcattta aatatatata tatatatata tatatttttt 7380ggctcctgat tctcttccgt cttcccatgt ggctgcattt taaaaggctt ccctaagatc 7440gttacgatta aatcaaccct ccccaggcat ctttaccgag ggctgtggtc cccaaagcga 7500tacagcccag gagggagaga ggctttggtg acttggagga aggactgtgt ccctccttag 7560ggcgtctgtg gcctcagtga gggaaggaag ctgcatcaga caggggtttc ctcgctgtcc 7620acccctctgg cagaagatgg attgggctgc cccgtataaa ttaatgaaaa gattaaagtt 7680tcgctaaagg ggacatcgag tttatgtgtc atctcctggt gtctgtgtgc ctgggatctg 7740caatatatcc cagcccttga tgtactgttt ctataaaaat aaattacttg taatttaatt 7800ccacactatt tctttccgta gtctattacc gacgagagca cgttagttca gctgcggaaa 7860attggttgtg gggtgtgtgc ggaccccgag aacgccctaa aataaagaca aatcggggac 7920aagctggggg ttatcgattg caggggtcgc atgaaaattt aacgacggta aataataata 7980aaaacaaaca tgggaatgca ataaaagaca taattctcca tcgccgcggg gggaaaggat 8040cctatagtaa aggcgagtgc gctttgaggg gtcataaaaa tcaattagtt ccaacaccca 8100cgtcccgcgt tgaggggacg gggacgagca gggacagaaa aagaaaccat atttgaatcc 8160catctctctg tgaattcttg ggtcacatgc gtctcagtac agcccgtccc gtgctgtgac 8220cggatagagt ttcaatttac tgtggaaatt tgctgtaaat aaattgagca tccgatagaa 8280gctgttgctg attaaccttt tatttttagc gtggccctgc aaagtcgtat cacccagctg 8340tcaggcttct aatcgaaagt tatgagacca cggtgagggg caggcggtaa tttaattaca 8400acaaatatct ttgggtttat ggcgcagagc taaattaaat gtcattattc actgtctgta 8460atggaaatca aaaggaaatc gcattacggc atttgggaaa gaaagcgggg agtgctcttt 8520aatgaagaaa taactgtctt aagcagtgtc acacacttca cttaccatat tcgggcctaa 8580ttggaatgga tcgtgaatca ctccaagact gatttattag cgcttcacgc agcggctaat 8640tcatcacttg tattcttcat catttttttt tttcctctcg ccgtgttgaa gggagagtga 8700atgaggcttt ccacgtttca ggaggatttt cttttttgaa aaatgccctt ccagaggctt 8760ttgggtggct ggcttgcttt ctgggccctg gaggagacag gcggagagtc caggtgggca 8820tggagaggca cagtggcagg tcacctggat ggtcagtgga ggtggaggtc tgaaggcgcc 8880agctttggaa attattggtg aatttcgatg tcagcaccag gcaggggcct ttttggcggg 8940ggtgtgaggg aggatgactt tgctgggaaa caggatcagg ttctccaggc gcactgcagc 9000ccggtaggac ccactttgga aatgaaaagc cagttccgaa agctgggctg gaagcttccg 9060tgttgggttc aagagcaagt tcacgttgcg ctgtgtagac tcctggctgc tcccaaactc 9120tgagggtttt ctgaggttcc cttcataggg gcaccggccc tgggccatgc acagtgcgta 9180agggtggctg tgggccgagg gacccagcac gtgttttgcc cacaacagcc ggagtgactg 9240gttcactcac cgccttggcg gaggacgcct gttctctgga cgaatcattt ctcttgggtg 9300gtgactgcct tgtgggtcaa ggtgcaggtt ttctgccaca gaaaacctgt taggaggaat 9360taagcgacta agactgtcag ggaggtggtg gtgggggaga ggagggggtg gtgtccagat 9420taccaggcat aggctaaact gcctgcactc tccagctggt ctgtctgtgg aggaggggat 9480tgtcaatact gggagagcag aggaggctcg taggaggtga gagggggtgg aatttgcatg 9540caaatcttca catgaggcct gtgtgaattt ctccagcctc ctgagggtcc cctgcgctat 9600tgcactcaac ttcttgatag tttaccccaa gactcagaag tccttagagg ggcagaatgc 9660ccccaccaca aagcctgcta tccttgggcg tcctcaggac ccttggtcat gaatgggacc 9720ctttcatgta tggggaccct tggtaatatg aatgggacgc cttcagctcc ccagggcttc 9780cgaggaggcc gagaagggca aagacacttc cgaggaggcc gagaagggca aagacatttt 9840ctgggcttgg tgtgtcaaga gctagattgg agaaggggct ggatttggaa ctctttagcc 9900atcagctcac cctctccgtt tgtggctaaa gtctgaaggt ggaaacttcg gttctcctac 9960agggtctaca ggagttgggg ggcggggcgc ccacacagaa cgctggaaag ttcgacagtc 10020cacttccact ggctcggaac tcactttttc accttaagtt catcagcggt aacgcatagg 10080tctcacttag gcagggcacg gatgatttaa caatttctac ttctaggtca ggtgcggtgg 10140ctcacacctc taatcccagc actttgggag gcccaggagg gtggatcgct tgaggtcagg 10200agtttgagac cagcctggcc aacatggtga aaccccgtct ctactaaaat acgaaaatta 10260gccaggcatg gtggtgagca cctgtaattc cagctactcg ggaggctgag gcaggagaat 10320cgcttgaacc tgggaggtgg acgttgcagt gaggtgagat cacaccactg cactccagcc 10380tggatgagag agcaagactc tgtctcaaaa acaaaataaa acaaaaacaa aacaaaaatc 10440aaaaaagaaa acccaatttc cagttctagg ccaggtgcag tggctcacgc ctgtcatccc 10500agcactttgg gaggcccagg agggtggatc gcttgaggtc aggagttcga gaccagcctg 10560gccaacatgg tgaaacccca tctctactaa aaatacaaac gttagctggg tgtggtggtg 10620tgcgcctgta atcccagcta ctcgggaagc tgaggctgga gaattgcttg aatctgggag 10680gtggaggttg cagggaggcg agatagtgcc actgcagtcc agcctggacc agagagcaag 10740actccgtctc aaaaacaaaa gaaagcaaaa acaaaaaaca agagaccagc ctggccaaca 10800tggtgaaacc gcgtctctac taaaatacaa aattagccgg gcatggtggt gggcacctgt 10860agtcccagct actcgggagg ctgaggcagg agaatggctt gaacctggga ggtggagctt 10920gcagtgagcc gagatagtgc cactgcactc cagcctgggc gacagagcga gacttgattt 10980cagaaccacc accaccacaa caaaacaaaa caaaaaatcc aaaaaaaccc caatttccag 11040tactaggtag tcagtgatgc agggctggag acagaggggc ggtaagtgtc tgggcgccca 11100ccatcagtca cctcccagct cccagaggtg caaagtgctt ggttcagcct catgggaagg 11160atgctccctg gggaggctgg gctgggttca cagggctctt cacatctctc tctgcttctc 11220cccaaggttt ggttccagaa ccggagagcc aagtgccgca aacaagagaa tcagatgcat 11280aaaggtgggt gtcgggactg gggggacctg aagctggggg atcctgctcc aggagggatg 11340gggtcgacga ggtgctggct acacccagga ccaccacact gacacctgct ccctttggac 11400acaggcgtca tcttgggcac agccaaccac ctagacgcct gccgagtggc accctacgtc 11460aacatgggag ccttacggat gcctttccaa caggtagctc actttttctt cctctgaaga 11520tccctaggga cctgctgctc ccttcccctt tcccctattt gctgccgcat cctgacactc 11580ctagtccctc cctgcccctg cagacttctc agctggccct tagaaaaaaa gcctcttttc 11640cgaggaggca tttacaggca ccttggcacc tatgaaatca ggctgggcca ggcggggtgg 11700ctcacacctg tcatcccagc actttgggag gctgaggagg gtgcatcacc tgagatcagg 11760agttcaagac cagcctggcc aacttaacga aaccccgtct attaaaaata caaaatgggt 11820gtggtggctc acgcctgtca tcccagcact ttgggaggcc gaggcaggtg gatcacctga 11880ggtcaggaat tcgagaccag cctgaccaac atgctgaaac cccgtctcta ctgaaaacac 11940aaagcttagc cgggcgtggt ggtgcacacc tgtgatccca ggtacttggg agggagaatc 12000acttgaacct gggaggtgga ggttgccgtg agccaatatc gcgccactgc actccactct 12060gggtgacaga gtgagactcc aagactccat ctcaaaaaaa aaaaaaaaaa tcaggctgta 12120aaaatccact tttgggaagg tgaacacaca caagcccaaa cagaaatctg acaaaaacca 12180gaggggtgaa aagtccacac agtcaggcac ccccacctgg cttgctgcct ggttaagaag 12240ggcgcagatg cctgtgcctg gataccagag atgggacaga cacccattcc cttttcatca 12300ccacccccga gtgcccgagg gcctggggcg tctgcctggc ccctggcccc tggcttgggc 12360tctgcacctc tgaactggag acaccctact cagctcccca cttactttgg agtgagcagc 12420gcttgggtgc ccagcgtgga tttggggctt ccagggagtc ggggttcggt cgcggagccc 12480aagcttccca agggcgcccc cgccctgccc tggcttagtg gtggggatgg gatgggggga 12540aacggggagc tgcgtggaag gaggtgaagg gtcacaggag gagagagcgc agcgcccacg 12600tgcgccctgc ctgaacgcgc agcgcagcgc ccggctgcgg tgccccttgc cccttcggtc 12660cctaatttgg ggatcgggag tgcatgcgcg ggcggaacgg gcttgggggg ggggctctgg 12720cagggcggac gcgtggcctc ccttcttcac cgttttattc caaggggaca ggctggggat 12780tgtatttggg cgcgtgtttg gctgagggtg cagggacttg gggggtggcg gtggggagcg 12840cggaaggtat aaacgtataa atcataagta aacaactcag aaatggaccc cgagcgctgg 12900tcgccgctag ctctccagct ctccctggcc caggcccgaa ggagaggggt ccgcatccct 12960ccgcggttct cctctcctgg gtacctggcc ttgaggtggg ggaacgagcc tacttcttgt 13020accgtctttt gccgacggcg ggacccagtg aaattaggcc gttggagccc gcaggcctgc 13080ctggctttgc gcaccggagt cttggggacc tggtgtcccc gggaaaaact tggggacctg 13140gtatccccgg gagaggcttg gggacctggt gtcccgggag aggcttgggt acctggtttc 13200tctggaagag gcttggacac ctggtgtcct gggagggcct ttgggacctg gtgtcctggg 13260agaggcttgg agatctgttg tcctgggaga ggcttgggga cctggtgtcc ctggagaggc 13320ttggggacct ggtgaccttg gagaggcttg gagacctggt gttctgggag aggcttgggg 13380acctggtgtt ctgggagagg cttggggacc tggtgtctct ggaagaggct tggacacctg 13440gtgacccggg agggccttgg ggatctggtg tcccgggaga gccttgggga cctggtgtcc 13500tgggagaggc ttggggacct ggtgaccttg gagaggcttg gggacctggt gtcctgagag 13560agccttgggg atctggtgtc ccaggagagg cttggggacc tggtgtctct ggaagaggct 13620tggacacctg gtgtcctggg gagaggcttg gggacctggt gtcctgggag aggcttgggg 13680acctggtgtc ctgggagagg cttggagatc tggtgagccg ggagaggctt ggggacctgg 13740tgtcccggga gaggcttggg gacttggtgt cccgggagag gcttgaacac ctggtgtccc 13800aggagaggct tggggacctg gtgaccttgg agaggcctgg ggacctggtg acccgggaga 13860gccttgggga cctggtgtcc tggggagagc cttggggacc tggtgacctt ggagaggctt 13920ggggacctgg tgtctcggga gtgccttggg gacctagtga cccgggagag gcttggggac 13980ctggtgtccc gggagaggct tggggacctg gtgtcctggg agagccttgg ggatctggtg 14040tcctggggag aggctggggg acctggtgtc tcgggagaga gccttgggga cctggtgacc 14100cgggagaggc ttggacacct ggtgtcccgg gagaggcttg gggacctggt gacccgggag 14160agccttgggg acctggtgtc ctggggagag gctgggggac ctggtgtctc gggagagagc 14220cttggggacc tggtgacccg ggagaggctt ggacacctgg tgtcccggga gaggcttggg 14280agcctggtgt cccgggagag ccttggggac caggtgacct tggagaggct tggggacctg 14340gtgatcttgg agaggcttgg ggacctggtg tctcgggaga ggttacgggg gctggttggg 14400ggagagaacg ttgtgagcca aagtccctga atccctgcga aaagagcgca tcgggagctc 14460cccctgaggg cgttccattt gtggaccccc ctcccatgcg ctttgcaggg agctgttcgg 14520attcccctgg cccggctccc gcggatgcat ccagtggcag cgccaattct gggccagggg 14580gaaggaggaa aggcgggtgt ggggtggtct ccacggctgg agaaggggcg acgctcccta 14640ggggagaaga ggcacgttgg gggtttccgg gggcgcgggg cggagcaggc cccccagtcc 14700ccatcctgcg ccctcacccc gccgggtccg ctcccgcagg tccaggctca gctgcagctg 14760gaaggcgtgg cccacgcgca cccgcacctg cacccgcacc tggcggcgca cgcgccctac 14820ctgatgttcc ccccgccgcc cttcgggctg cccatcgcgt cgctggccga gtccgcctcg 14880gccgccgccg tggtcgccgc cgccgccaaa agcaacagca agaattccag catcgccgac 14940ctgcggctca aggcgcggaa gcacgcggag gccctggggc tctgacccgc cgcgcagccc 15000cccgcgcgcc cggactcccg ggctccgcgc accccgcctg caccgcgcgt cctgcactca 15060accccgcctg gagctccttc cgcggccacc gtgctccggg caccccggga gctcctgcaa 15120gaggcctgag gagggaggct cccgggaccg tccacgcacg acccagccag accctcgcgg 15180agatggtgca gaaggcggag cgggtgagcg gccgtgcgtc cagcccgggc ctctccaagg 15240ctgcccgtgc gtcctgggac cctggagaag

ggtaaacccc cgcctggctg cgtcttcctc 15300tgctataccc tatgcatgcg gttaactaca cacgtttgga agatccttag agtctattga 15360aactgcaaag atcccggagc tggtctccga tgaaaatgcc atttcttcgt tgccaacgat 15420tttctttact accatgctcc ttccttcatc ccgagaggct gcggaacggg tgtggatttg 15480aatgtggact tcggaatccc aggaggcagg ggccgggctc tcctccaccg ctcccccgga 15540gcctcccagg cagcaataag gaaatagttc tctggctgag gctgaggacg tgaaccgcgg 15600gctttggaaa gggaggggag ggagacccga acctcccacg ttgggactcc cacgttccgg 15660ggacctgaat gaggaccgac tttataactt ttccagtgtt tgattcccaa attgggtctg 15720gttttgtttt ggattggtat tttttttttt tttttttttt gctgtgttac aggattcaga 15780cgcaaaagac ttgcataaga gacggacgcg tggttgcaag gtgtcatact gatatgcagc 15840attaacttta ctgacatgga gtgaagtgca atattataaa tattatagat taaaaaaaaa 15900atagccgtgc actcttgacc ccgtcaacgt ccaacgtgga aaaggcgtta cctcttctcc 15960cagcgctggc cgcctggcca ctgagggccc tttgcaaaaa tcacgggtgt agagatggcc 16020ctgggcgcgc tgggagtgtg gttgtgtttc tgaaggggat aaaagagggc acggtggtgc 16080caagatatca gtttggtacc tgagctgttt ctggttggga agcgtaaaag ccagggagag 16140atccagagag ttttcaagtt tttgcagatg taggtggttc cagcttttct ttctccccta 16200ctccatcttc tgcgttcccc cagttctttt atttctttgt tttttatttt tgagacagag 16260acttgctttg tcgcccaggc tggagtgcag tggcgcaatg tcagctcact gccacctcca 16320cctcccgggt tcaagcgatg ctcctgcctc agcctcccga gtagctggga ctacaggcac 16380ctgccaccac ccccggctaa ttttttgtat ttatagtaga gacggggttt caccgtgttg 16440gccaggctcg tctcgaactc ctgacctcag gtgatctgcc cgcctcggcc tcccaacgtg 16500cccccagttt tataaacagc agatagcaac ttgtcgtcac agctggcatg ggctggacag 16560ttgcttgaaa tgacctaacc aaaaacattc aagggttctg cccccagatt tcgggagatc 16620cacgttccat gttctgattg gttttctggg aacacagcaa ggggtttggt gacctccgag 16680aagatccatc tgcatgattg gcattagtta ccacagcctg cccagagaga aactatcttc 16740tcccaacatt tactaacatc cactggtcaa ctctcttatt tccataacac atttgcatct 16800ttctggattc aagcttggtg gttttctttc ctaacttctg atttagatac ttctccctga 16860ggtggggata aaagaaaaaa aaaaaacaac ttcttttttt cttccgcata acactttcta 16920tcttgtcact gagctgaact gtagatccat ttggacccgt ctcatttgta tcttctgata 16980ttctttatac aaaccaaaag tccccttcaa cattttttat gtcaaaatgt tacaaccgct 17040gtaaaatgac ggagagagag agaaagaatc ccagacatta acggtattag agagtttgcc 17100tcattcatcc atttttctta aaagctggaa attaaaaaaa aaaaagagag agagaggctt 17160taatagttaa gctgaaattt ttatcgaaaa gaagaattgc attttgaatc tttgggaagt 17220aggttcattc atcagagtat gtaacccttt ggaaaagtgg ttggtaagat atgtacagcc 17280ctagattttt ttttttttaa ccaaaaaggc tgagtaattt tgaaaaatcg aaacataaca 17340gtgtgtcatc atttcctccc aagaaaaagc tcactccacg tgagtagaaa gacatctacc 17400tggtccctgt agaatctgaa cgtttctctt tagagacgga atttcaatct tgttgcccag 17460gctggagtgc agtggcacaa tctcggctca ccgcaacctc cgcctcccgg gttcaagcca 17520ttctcctgcc tcagtctccc gagtagctgg gattacaggc acctgccacc aggcctgggt 17580aactttctgg tatttttagt agagacaggg tttcagcctc ccgagtagct gggattacag 17640gcacctgcca ccaggcctgg gtaactttct ggtattttta gtagagacag ggtttcagcc 17700tcccgagtag ctgggattac aggcacctgc caccaggcct gggtaacttt ctggtagttt 17760tagtagagac agggtttcgg cctcccgagt agctgggatt acaggcacct gccaccaggc 17820ctgggtaact ttctggtatt tttagtagag acagggtttc ggcctcccga gtagctggga 17880ttacaggcac ctgccaccag gcctgggtaa ctttctggta tttttagtac agacagggtt 17940tcggcctcct gagtagctgg gattacaggc acctgccacc aggcctgggt aactttctgg 18000tagttttagt agagacaggg tttcagcctc ccgagtagct gggattacag gcacctgcca 18060ccaggcctgg gtaatttttt tgcatttttg gtagagacag gtttttgccg tgttggcccg 18120gctggtctca aactcctgac ctcaggttga cctgcccgct ttgtccctcg caaagtgctg 18180ggattacagg cgtgagccac cacacctggc ctgaatctga acttttaaaa gggagttact 18240gactctcaac tgtgcgggga cggtttcact ttgatttaat atggaaagag ggccaagtgt 18300catcctcaca aatgggtccc cgaagcagat caaacgcaga gaactgtgag ggtgggacac 18360gagtgtctgt ggacactggc tgcctttggc ttttctcctg cgagagaagt tgggtgactt 18420tctgtaggtg gatgagtgat ccctgaatga gtgtggggta cgtgtatgct agctgcttct 18480ttctccctga aactctcgga tggaaggaag taagaaattc agcttgggct gtgaccagtt 18540ctcaccacca acgccctctt ctctctccct tctccttcct tccttccttc cttcctttct 18600ttctttttct ttctttctct ctttctttct tttctttctt tctgtttctt tcctttttat 18660ctttctctct ttttctttct cttttccttt tttgtttctt tctttctttt tctttctttc 18720tttttctttc ttctttcttt cttcgatgaa gtctcactct gtcacccagg ctggagtgca 18780gtggtgcaat cccagctcac tgcatcctct acctcctggc ttcaagaaat tctcctgcct 18840cagcctccca agtagctggg atgacaggca cccaccacca ttcccggata atttttgtat 18900tttttagtag agactgggtt tcgccatgtt ggccaggctg gtcttgaact cctgacctca 18960catgatccac ccgcctcagc ctcccagagt gctgggatta cggggtgagg caccgcgccc 19020ggcctcctct ctctttttct gagatgttta ggaaggactg ggctgatggg gaccctctgt 19080atgtgatgtg cgtgggtttg gtttcccgga aggccctcca gagacacgtt tgcgtgaaca 19140ttcagcatgg aaacaacata cgtctctcca caggaggtga gaaattgaat ttatggggtg 19200ggtgtacgct ggcgattctt ggtgcttttt gctcaaaaca aggttctttt gaaagtcacg 19260ttcctgcttt ccctgtggct tcccggtgag ctcgctcgca gagcaaggaa taccacccag 19320agagcaacgt gggctgtgtt ccgttgtaac gccgttgcag agagaggatt tggtgtgtga 19380gatccgtacc agctccagca cactgatagg aacacgttgc tggccgaact gaacgatgct 19440gggttgggtc ctgattgata cgtattttct tccctcctct ccccaaaact tggccaaata 19500gtccgtggag ggttgtcagt cgccgcagtt gagcaaaaaa cacttcttcc tttgagtggc 19560tgttctggtg aaatctgttt ctgacatatc cacttttctc tctcttttct ctctctctga 19620ctgcgaagca cccacaggga gaaggaattg gatgtatcgg atgttggtat tagattttct 19680ttctccgttc gagtctctga ctggtgcata ctttgcaaag gtgtgttcct ggcaattgcc 19740aagagttaga aaaatgcacc ttctctggtg gccgttgggg tgttgtttca caggcagtgg 19800tgacagggcc ccttggctgt ggctgtcttc tccagcgccg tggataaaga gacgggacag 19860attctgtgcc tctgtacgat ttagagcgta actgaccgcg tccaacaccc gtttttccac 19920ttacaaagct ggtggtgcga cgggcttggt gtctcccgta cgggaaggag gcctttgggc 19980cgctccaaag acgccctgtc gtaggaatgg cctctccatc ccgccaaagt ccagccaggc 20040ccccgaaatg gtcccatttc cttggaagcc tgagtttctg ttctggtctt gctgctgtcc 20100ttggccacgt cagcacgtgg gagcatctgt ggataccgca gagtctgggg acagctgggc 20160gtttaaccga aatgaagccg agacgggttt caggttttgg tgccaagctc tggtcaggat 20220gaaagggaaa taccagagtc ctctgtcctc gcctctgggt ttcatgctga cctttctaac 20280atttgttttc ccctaagaac aagcagaagc ctccagctcc ctttagctcc acagttttcc 20340cggggacata gcgaggatgg cacacggcag ccactcccac gacacacatt tcggaggcac 20400tttgctggaa gccgcttgtc tcctccagct ttgggaggtc tggggaggag agaggctttc 20460ggtggacacg tttgacatta aaaaaaaaaa aaaaaaaaaa aaaaaaactg gtgcctaatt 20520tattaaagag aattagctta gcgagtatat gctgatattc ttcgacacac gtgggtaagt 20580tgatgccatt tataaatgtt ttattgaaat ttgatattta atgagaagcc ggttaaggaa 20640tgtagacaat atcccgtttc aaagctatga aatgtgctat ttattgaaag gggatgtggc 20700ttcacgagtt cagcccattg tacgtgcagg tcccgtggga aggaggcaaa agcccctgct 20760tcttactttg tgatgtatgt gcatttgtta tttatttttt tttccttggt cggacgttca 20820taaatatgta ctattttaat tatgtcgagt gtaaatttga catcgcgttg catttatttt 20880tatatttctg aaaactgttg ctttttcttt ttccctcccc cattgacgac atagcggccc 20940ccgcgtccgg gttacaaata catctacaga tattttcagg gattgcttca gatgaaaaca 21000aatcacacac cgtttcccaa accaacagtc ttcacatttc tatccctctg ttattgtcgg 21060caggcggtga ggggtagaaa aaaaacaaac aaacaaacag aaaaaaaaac caaaaaaaac 21120caccctgagt ttctctggtg acgccctcat tctcctaacg ttcaataatc tcaatgttga 21180gttgcagcaa cagactgtat ttttgtgacg ccccgtagta tgaatgtaca tcttgtaaaa 21240ctgagatata aataaactta taaatatttg tattcaagtg ttaaaaaaaa aaaaattctc 21300aacctctccc ctgaggacag gcttattgga aaaaaaaaaa aaaaaaaaaa atcctgagtc 21360ggccgtggct gaacacagag tgttgttctg ctccgtgcat ttccagggtg ggtacccagt 21420gttgcccccc agccttagat cgggaggtac cattgacttt tgcttgtatc ccatcccctt 21480cctttactga aacctacctc cccgcttctc agccaacgtc cccccagaag gtggcaaaaa 21540aaacagagga aaaagccctg atttgaatca agtcagagct gctaattctc cactttcttt 21600aattaattaa tttatttttt tttttgagac tgagtctcgc tctgtcgccc aggccggagg 21660agtgcagggg cgcgatctcg gctcaccgcg acctccgcct cccgggttca agcgactctc 21720ctgcctcagc ctcccgagta gctgggatga cagtcacctg caccaccgcg cccggctcat 21780ttttgtattt ttagtagcaa tggggtttca ccgtgttggt caggctggtc tcgaactcct 21840gacctcgtga tccacccgcg tctgggcccg gccggtgatg tgtgtgcttt taacttttat 21900tttgttccag ttttcgacag tggcacggat tttccagcac ggtcttgcaa ggatgattga 21960gtcatttttg agacaaaaaa tataataata ataaatggaa aaagaaatcg acttttaaaa 22020atgacaaatt tttttttttt ttttttgcat agatttttct ctctttatgt aaaggaaagt 22080tcatgattgg atttggccgg cctgactgct tcccggctgt gataaaaaac acatgtgagc 22140tgggagggaa gtgggggagg gacacagctg cccacacagg gttcccaccg cggttacagg 22200gtgggcagtg ctgggggagc tttctctgtg gggggctcag agcctgagga caggtgagcc 22260tctccgacac ctccccagtt gcctggagtc taaaccgtcc gttgtctgta ccgtccgttc 22320ttcctgctga ctcctggtag ttcctgaaag cttctcttgg ccagagaagg ggtttcagag 22380gccgtgtgtc caggccattc tgcaaagtgc aacttgaccg ttcctttcct tttctggcct 22440gcgtggtctg aagctcagag ccctctcttc acccagcctg tgtgtgtctt gccggacaga 22500agaaaaatgg tgctttttgc gtgttagcag aggtgctttt catggctgac ctcaacgcgt 22560ccatctccag ccttgaccaa gctgtttttt aggggcaaac gcaggcaagt tctgaatgca 22620cacagttatt tcatggttaa actattcagc tttggccggg cgcagtgtgg ctctcacgcc 22680tgtcatccca gcactttggg aggccgaggc gggtggatca cctgaggtca ggagttcgag 22740accagcctgg ccaacacggt gaaactctat ctctactaaa aatacaaaaa ttagccgggc 22800gtggtggtgt gtatctgtaa tcccagctac tcaggaagct gaggcaggag aatcgcttgg 22860acccaggagg cggaggttgc actgagccga gatcgcgcca ttgcactcca gcctgggcga 22920cagagccaga cgctgtctca aaaaaatgaa taataaaata aaataacagg aactaaataa 22980aataaaacgt tcagctttgt tctgcaaatc cactcctatt gttttacgtg gtttgagaga 23040ctctgtccct tagaaataga tgtttgttgc caattgtaat gaatctgttt caaaaatgaa 23100cagaatattc aaatggtttg agagatcttt tcccttagaa atagcttgtt gccaatcaca 23160aagaatgttt ttcaaaaatg aatggaatct tcctggatat cgcttccaga tcttcatttt 23220ttttgcatag ttcaacctga aaagtaagtg tctcagccct gaatttcttt ctgatttttc 23280catgggttgt cttgcagact tctctggact tgaccacatt taaaaaaaaa aaaattaact 23340ttttcacacg gacacggttt caataggaat gagatctttg agtttttatg taacagattc 23400ttaccatcag ttctcagatt cccaaattac acacaaaaag ccacggactt cgcctcctgc 23460taacatgtcc ttctgtttct gaggcttctg ttggtgttag actttcatgt ttaatagcag 23520acaatgtagg gatttaaaga aaaatgcaga gaaagcaaaa acactgacca aacacacgga 23580gataagcttt ctaaagcctt tgttcttgga gttgtcgtta aaaaaaaaaa gttgttttaa 23640actttgcaag catgcctata ttgaactcat aagcaagaga gccaagaaaa atagtgtcgg 23700tcgtctactc tacacgtttt cccaaaacag acgtatttta atttcttttg tttgaactca 23760cagatgctga gagttaaaag ttaaattttt gtcatgaaca atagtggcca aaaccacagt 23820tacttttgca ctatagcata ataagaaaaa tacaggctgg gctcggtggc tcacacctgt 23880aatcaaagca cttttggagg cgaaacagcc agatcccttg agcccaggag attgagacca 23940gcctgggcaa catagcgaga ccctcatctc tacaaaaaag gtttgttaca tatgtaacaa 24000acctgcacat tgtgcacatg taccctaaaa cttaaagtat aataataaaa aaattaaaaa 24060aaaattcacc aatcaactgc ctgctggtgc cttcaagaga ctcacctaac acataaggac 24120ttgcataaac ttataaaaca attcaatgga agaatccttg aaagtattct gagaagacag 24180tataataaac tgatttctaa aaaggctata aaaaattgaa taaatcattg ttgggcatcc 24240tgtgctgaaa tataatgcag ccaataaaaa ttacaaaatg aataaacatt ttataacaat 24300aaaaaaaagt caaataatta ggcaggcatg gtggtgctct cctacggttg aagctattca 24360gcaggcaaga ggatactttg tttttgtttt ttaatttttt ttgagacaga gtctcgctct 24420gttgccaggc tggagtgcag tggcgtgatc tcagctcact gtaatttctg cctcccgggt 24480tcaagcgatt ttcctgcccc agcctcccga gtagctggga ttacaggtgc ccgccaccac 24540acctggctaa tttcttttgt atttttagta gagacgaggt ttccccatgt tggccaggct 24600ggttttgagc tcccgacctc gggtgatcca cccgcctcag cctcccaaag tgctgggatg 24660acaggcgtga gccaccgcgc ctggcccagg aggattattt gatcccagga ggtggaggct 24720gcaggaagcc atgattgcac cactgcactc cagcctggct gacagagtga gaccacatct 24780ctaaataaat gaataaatac aggcagaaac tttttttgtt ttgttttgat ggagtcttgc 24840tctgtcacca ggcaggagtg cagtggtgcc atctcagctc actgcaacct ccacctcctg 24900ggttcaagca atcctcctgc ctcagcctcc cgagtagctg ggattacagg tgcccgccac 24960cacgcccggc taattttttg tatgtttagt agagacggga tttcaccgtg ttagccagga 25020tggtcttgat ctcttgactt tgtgatctgc ctgcctcagc ctcccaaagt gctgggatta 25080caggcatgag cccaggagtt caagaccagc ctcagcaaca aagtgagacc ttttctctcc 25140aaaaaatcaa aaatttagcc agctgtggtg gctcctgccc gtgatcccag tactgtggga 25200ggctgaggca gaattgcttg agcccaggag ttcgagacca acctcagcaa aaaggactct 25260ctctctctct ctctctctct ctctctctct ctctctatat atatatatat atatatatat 25320gagtttcaaa aattgctggg tgaccagctc atctactggt tttccccttg ggaaagtgaa 25380attgtcatgt attgaagatt tccaaggaag ttgtattgaa tgagaaacaa actcaatctg 25440ttcgtgttta aagagctgca gtgcgtttgc tgtgtttccc ataaaactgc acttccaaaa 25500gacacgctga gaaaggagac caggatttgt aattcagaaa ttggaaagca agttaggctg 25560gacgtggtag ctcatgcttg ttgtaatctc agcactctgg gaggctgagg caggaggatc 25620acttgagccc aggagttcaa gaccagcccg tgccacatgg tgaaaccctg tctctccaaa 25680aaataaaaca tttagccaga tgtggtgact catgcctgta atcccggtat tctgggaggc 25740tgaggcagag ttgcttgagc ccaggagttc aagaccagcc tcggcaacaa agtgagaccc 25800tgtctctcca aaaaataaaa catttagcca gctgtggtga ctcatgcctg taatctcagt 25860actctgggag gctggggcag aatggcttga gcccaggagt tcgagaccaa cctcagcaac 25920aaagtgagat cttgtttctc caaaaaatca aaaatttagc cagctgtgct ggctcatgcc 25980tgtaatcccg gtactctggg aggctgaggc agaatcgttt gagcccagga gttcgagacc 26040aacctcagca acaaagtgag atcttgtttc tccaaaaaaa tcaaaaattt agccagctgt 26100gctggctggt gcctgtaatc ccggtactct gggaggctga ggcggaattg cttgagccca 26160ggagttcaag accagcctca gcaacaaagt gagatcttgt ttctccaaaa aataaaacat 26220ttagtcagct gtggtggctc aagcctgtga tcccagcatt ttgggaggcc gaggcgggcg 26280gatcacgagg tcatgagatc gagaccatcc tggctaacac ggtgaaaccc cgtctctact 26340aaaaatacaa agaaaattag ccgggcgtgg tggcgggcgc ctgtagtccc agctactcag 26400gaggctgagg caggagaatg ccgtgagcct gggaggcgga ccatgcagtg agtcaagatc 26460gcgccactgc cctccagcct gggccacaga gcaagactcc gtctcaaaaa aaaaaaaaaa 26520aaaactgctg cccaacctgt gtttgcacca ctgccctcca gcctgggcaa cagagcaaga 26580ctccgtctca aaaaaaaaaa aatgctgccc aagctgtgtt tgcaccactg ccctccagcc 26640tgggcaacag agcaagactc cgtctcaaaa aaaaaaaaaa aaaatgctgc ccaagctgtg 26700tttgcaccac tgccctccag cctgggcaac agagcaagac tctgtctcaa aaaaaaaaaa 26760aatgctgccc aagctgtgtt tgcaccactg ccctccggcc tgggcaacag agcaagactc 26820cgtctcaaaa aaaaaaaaaa aatgctgccc aagctgtgtt tgcaccactg ccctccagcc 26880tgggcaacaa agcaagcctc agctttctgc catctccaca accaagaaag caattcacac 26940agaaatcagt gcatcgtgca gtgacctctt cagaaaacca atgagttttc cacctgagga 27000actgtttctg agccccattc agaaaaacac atccctgtaa ctgcagggca gatttactca 27060ctgtatgcct gtttaaataa agcttccagc ctctgcatgg ggtctgtctg gaagctcctg 27120tatctgtccc acattcttgg aatcacaatg cacccttggg aggaagatat gtatttaaag 27180ggagtggatg ttatggtgag aaaatgctgc ccatccttct agaagacaaa agccacacaa 27240aatacatcac aagaaccagt ttttttcaga gaagaacctg cacaaagaac ctgctccccc 27300cacaccccca cacacaggtg aattaacagg atgtatgttt tatcataaaa gcacaggttt 27360gtttcctatg cactctctga ggatttggcc atatgcaaag atgtacaaaa accttctctt 27420tccccaggga accgtaaccc gtctgaaaag atgcccttct cagaagcgag ttgaacgatt 27480gttggaaaag ataaaatacg acgtgcacac acacagtaga gaaatgtcac ccatgcaaat 27540tatgtgtttg aatggaacac attcaggaag ctaaatgggg tatgaccaca catttgggtt 27600gatttatttg acgagtggaa ggggcagatg gaaatgaata ctgctgtttt cctttggaag 27660gccatatatg ggaataccaa gaggattact ttggaagttt agcttctcca ggtggtctct 27720ctctctctct ctttttttga gacagagtct cactctgtca cccaggctgc agtgcaatgg 27780cgtgctctcg gctcactgca acctcagcct cccaggtaca agcgattctc ctgcctcagc 27840ctcccgagta gctgggatca caggtgtgca ccaccacgcc tggctaatgt ttgtattttc 27900agtagagatg aggttttacc atgttggcca ggctggtctt gaactcctga cctcaggtga 27960tccgcctgcc tcggcctccc aaagtgctgg gatgacagac atgagctagc acgcccggcc 28020ccaggtggtc tttttagcgg gtattaaagc agctttctct ctgagcctta aaccatgaag 28080atagacagac tcagtgtatg ggttttagag ttgtaatttt ataaaaataa gaaaaagtcg 28140acctatcatt gatggttagt attttttgta gcagttgcat gcaatattag gataaggcat 28200gttctcaaaa agaactcttt tttttttttt tttgagacgg agtctcgctc tgtcacccag 28260gctggagtgc agtggcacga tctccgctca ctgcaagctc ctcttcccgg gttcacgcca 28320ttctcctgcc tcagcctccc cagtagctgg gactacaggc gcccgccacc acgcccggct 28380aattttttgt atttttagta gagacggggt ttcaccatgt tagccaggaa ggtctcgatc 28440tcctgacctc atgatccgtc cgcctcagcc tcccaaagtg ctgggactac aggcgtgagc 28500cactgcactt ggcctttttt tttttttaga tggagttttg ctcttgtcgc ccaggctgga 28560gtataatggc atgatctcga ctcactgcaa cctccgcctc ccgagttcaa gcgattctcc 28620tgcctcagcc tcccgagtag ctgggattac aggtgcccac caccatgtca agataatgtt 28680tgtattttca gtagagatgg ggtttgacca tgttggccag gctggtctcg aactcctgac 28740ctcaggtgat ccacccgcct tagcctccca aagtgctggg atgacaggcg tgagcccctg 28800cgcccggcct ttgtaacttt atttttaatt tttttttttt tttaagaaag acagagtctt 28860gctctgtcac ccaggctgga gcacactggt gcgatcatag ctcactgcag cctcaaactc 28920ctgggctcaa gcaatcctcc cacctcagcc tcctgagtag ctgggactac aggcacccac 28980caccacaccc agctaatttt tttgattttt actagagacg ggatcttgct ttgctgctga 29040ggctggtctt gagctcctga gctccaaaga tcctctcacc tccacctccc aaagtgttag 29100aattacaagc atgaaccact gcccgtggtc tccaaaaaaa ggactgttac gtggatgttc 29160tagcttcctg ttctcgtctt ttctttgtta attgtacagt ttgagggtgt gtgtgcgtgt 29220gcgcacgtgt gtgtgtgcag tctcctgatt tcatgtattt aattgttatt accaccacct 29280ccatctctca ttccttctta ccctcactgt gtaaagatac atgttgtttt taaattttat 29340gtatttatat ttatttattt gtatttctga gacagagtct cactctgttg cccaggctag 29400tggcatgatc tcagctcaca gcaacctttg cctcctgggt tcaagcgatt ctcctgcctc 29460agcctcccga gtagctgaga ttacaggcac acaccaccac acccggctag ttttgttttg 29520agacggagtc tcgctctgtt gcaggctgca gtgcagtggc gtgatcctgg ctcactgcaa 29580cctctgcctc ctggattcaa gcgattctcc tgcctcagcc tcccaagtag ctgggattac 29640aggcgcccac cgccacacct ggctaatttt ttattggtag tagagacggg gtttctccat 29700gttgaccaga ctggtcttga actcccaacc tcgggtgatc cacccacctg ggcctcccaa 29760agtgctggga tgacaggcga gggccaccgc gtccagcctt cttcttcttc ttcttttttt 29820tttttttaag atggagtttc actctgttgc ccaggctgga gtgcagtggt gcaatctcgg 29880ctccctgcaa cctccacctc ccaggttcaa gaaattcttt tgcctcagcc tcccgagtag 29940ctgggactac aggtgcccgc caccacaccc acctaatgtt tgtatttttt tggtagagac 30000ggggcttcac cacattggcc aggctggtct tgaactcctg acttcagatg atcctcctgc 30060ctcagcctcc cagagtgttg ggattacagg cgtgagccac ggtgcccggc cagacgtcat 30120gtcttaggaa atcagaaagt gggtagtttc cgcactctga ggagaaaaag agacgtccgg 30180cgaagagaaa ggagagtgaa aggatgtctc ctcttgtctg tagcctgttc tcaatcgtga 30240gtgagccaat tgccagaaac tgagggtgct tcatttggcc aggcaagctt ctcaacagaa 30300tgtctaagta cttgttaatg ctgagaagct

ctccaagcta ctgcactcca gcctgggtga 30360cagagcacga ccttgtctga aaacaattaa ttaatcaatt aattaatata atgaaatcat 30420actgaactca ggagaccatt ggggtgggca gggctggggt tggaaaggaa cataaaatat 30480ggtgcaatgg actttgctcc agtctccctc cccatctctt ctcgccaaga gtctctggag 30540ggagcatggg gaagatgctt tgggaatctg taacttcttg tcttgtaaac agaatatcta 30600agtaattgtt aatgctgaga agttatagat ttccaaagcc tttctccagg ctacggacaa 30660gggtcatggg ttactcagtg ttacagaaag aatgacatgg agatgtttgt tacatcttaa 30720ggaaccatga ggggccagag tattttactc taagtgtaga tggtacattg gccacgcctg 30780tcccaacacc accaatggtg gcacctaact tttgtgtttg tgccccacat ttcttcttct 30840tttctgacgt aaatgcaagt gatattcctt ggaaaccatg ctgcagcaag aggccatctg 30900actactagtg ataccctgta gctcacctac agcagctcac ttgaagcagc tcacccatag 30960ctcaggtata gctcacctgc agcggctcac ctgtagctca cgtgtagctc acttgtagca 31020gctcactggt agctcacctg cagcagctca cctgtacctc acctgtacct cacctgcagc 31080agctcacctg tagctcacct gtacgtgagc caccgtaccc ggccagcaag accccatttc 31140taaaataaat acacaaaaat tagccggacg cggtggcgcg tgtctgtagt tgtagctact 31200caggaggctg aggtgggagg attgctggag gctgggaggt agaggctgca gtgaaccgtg 31260atccagccac tgtactctag cctggatgac atagcaaaac cttgtctcaa aaaacaaaaa 31320caaaaaacaa aacaaagaaa caaacaaaaa acccacacac accggaaaac aaaacaaaaa 31380gcaaaaagga aagaaaagag agccaggtcc caaatatata tttccttgga gaaccatttg 31440caaagagcac acttaaggcc gggcgcggtg gctcacgcct gtcatcccgg cactttggga 31500ggccgaggtg ggtggatcac gaggttggga gatcgagacc atcctggcca acatggcgaa 31560accccatctc tactaaaaat acaaaaaatc agccaggtgc tgaggcaggt gcctgtagtc 31620ccagccactc aggaggctga ggcaggagaa tggcatgaac ctgggaggtg gaggttgcag 31680tgagccgaga tcgcgcccct gcactccagc ctgggcgaca gagcgagact ccttctcaaa 31740taaataaata aataaataac aaagagcaaa cttaaaattg tctcagaaat cccacgggat 31800attggatctc cctcatgcct atctgatgac actttgagtg tctggggccc cgtgcctatt 31860ttctggggtt cccagaagct gccgttctga aagtgtggct ctcggggacg tggcacaggt 31920gtggatgtct gttttaaatg tcaggcgttt ggacgttgag gaacgtgagg ctgaaggtcg 31980ccttcgccga ccccctgagt ttagggtcct gccttttaaa atcttcccag cactctgttg 32040ttcacgcaag cgtcccatct gtttgggtgg ccgtgccgtc tgcatctgtc tcgaaccttc 32100acagctttgc agaatatcct gtttctcaat acggatggag aaacacgaga cgcgttttct 32160gggttatttt agccgtcacg gagaacccca gactcatgtg tgctaatgac ctcattaatg 32220atactctgag gcagacagcc ctgcctgatc ttaacaacat tttttaaatt tctttttttg 32280ttgttgttgt tacagcatca ttcatataac gtaggaaacc gtgatcagta gcttttagga 32340tatttgcaac agggtgtaac adaaabd 3236715806DNAHomo sapiensCDS(43)..(612)misc_feature(667)a, c, t, g, other or unknown 15gtgtccccgg agctgaaaga tcgcaaagag gatgcgaaag gg atg gag gac gaa 54 Met Glu Asp Glu 1ggc cag acc aaa atc aag cag agg cga agt cgg acc aat ttc acc ctg 102Gly Gln Thr Lys Ile Lys Gln Arg Arg Ser Arg Thr Asn Phe Thr Leu 5 10 15 20gaa caa ctc aat gag ctg gag agg ctt ttt gac gag acc cac tat ccc 150Glu Gln Leu Asn Glu Leu Glu Arg Leu Phe Asp Glu Thr His Tyr Pro 25 30 35gac gcc ttc atg cga gag gaa ctg agc cag cga ctg ggc ctg tcg gag 198Asp Ala Phe Met Arg Glu Glu Leu Ser Gln Arg Leu Gly Leu Ser Glu 40 45 50gcc cga gtg cag gtt tgg ttt caa aat cga aga gct aaa tgt aga aaa 246Ala Arg Val Gln Val Trp Phe Gln Asn Arg Arg Ala Lys Cys Arg Lys 55 60 65caa gaa aat caa ctc cat aaa ggt gtt ctc ata ggg gcc gcc agc cag 294Gln Glu Asn Gln Leu His Lys Gly Val Leu Ile Gly Ala Ala Ser Gln 70 75 80ttt gaa gct tgt aga gtc gca cct tat gtc aac gta ggt gct tta agg 342Phe Glu Ala Cys Arg Val Ala Pro Tyr Val Asn Val Gly Ala Leu Arg 85 90 95 100atg cca ttt cag cag gtt cag gcg cag ctg cag ctg gac agc gct gtg 390Met Pro Phe Gln Gln Val Gln Ala Gln Leu Gln Leu Asp Ser Ala Val 105 110 115gcg cac gcg cac cac cac ctg cat ccg cac ctg gcc gcg cac gcg ccc 438Ala His Ala His His His Leu His Pro His Leu Ala Ala His Ala Pro 120 125 130tac atg atg ttc cca gca ccg ccc ttc gga ctg ccg ctc gcc acg ctg 486Tyr Met Met Phe Pro Ala Pro Pro Phe Gly Leu Pro Leu Ala Thr Leu 135 140 145gcc gcg gat tcg gct tcc gcc gcc tcg gta gtg gcg gcc gca gca gcc 534Ala Ala Asp Ser Ala Ser Ala Ala Ser Val Val Ala Ala Ala Ala Ala 150 155 160gcc aag acc acc agc aag gac tcc agc atc gcc gat ctc aga ctg aaa 582Ala Lys Thr Thr Ser Lys Asp Ser Ser Ile Ala Asp Leu Arg Leu Lys165 170 175 180gcc aaa aag cac gcc gca gcc ctg ggt ctg tgacvccaac gccagcacca 632Ala Lys Lys His Ala Ala Ala Leu Gly Leu 185 190atgtcgcgcc tgtcccgcgg cactcagcct gcasnccctn ddkanmcgtt rctyhtcmat 692tacactttgg gaccycgggd bagvcctttt nnagacttyv atkggscwcs ctggbccctb 752rkgavvactt gsghycgrga accgakhtgc ccabaygagg accrgtttgg akdg 80616190PRTHomo sapiens 16Met Glu Asp Glu Gly Gln Thr Lys Ile Lys Gln Arg Arg Ser Arg Thr 1 5 10 15Asn Phe Thr Leu Glu Gln Leu Asn Glu Leu Glu Arg Leu Phe Asp Glu 20 25 30Thr His Tyr Pro Asp Ala Phe Met Arg Glu Glu Leu Ser Gln Arg Leu 35 40 45Gly Leu Ser Glu Ala Arg Val Gln Val Trp Phe Gln Asn Arg Arg Ala 50 55 60Lys Cys Arg Lys Gln Glu Asn Gln Leu His Lys Gly Val Leu Ile Gly 65 70 75 80Ala Ala Ser Gln Phe Glu Ala Cys Arg Val Ala Pro Tyr Val Asn Val 85 90 95Gly Ala Leu Arg Met Pro Phe Gln Gln Val Gln Ala Gln Leu Gln Leu 100 105 110Asp Ser Ala Val Ala His Ala His His His Leu His Pro His Leu Ala 115 120 125Ala His Ala Pro Tyr Met Met Phe Pro Ala Pro Pro Phe Gly Leu Pro 130 135 140Leu Ala Thr Leu Ala Ala Asp Ser Ala Ser Ala Ala Ser Val Val Ala145 150 155 160Ala Ala Ala Ala Ala Lys Thr Thr Ser Lys Asp Ser Ser Ile Ala Asp 165 170 175Leu Arg Leu Lys Ala Lys Lys His Ala Ala Ala Leu Gly Leu 180 185 1901718DNAArtificial SequenceDescription of Artificial Sequence synthetic primer 17gcacagccaa ccacctag 181821DNAArtificial SequenceDescription of Artificial Sequence synthetic primer 18tggaaaggca tcatccgtaa g 211926DNAArtificial SequenceDescription of Artificial Sequence synthetic primer 19atttccaatg gaaaggcgta aataac 262025DNAArtificial SequenceDescription of Artificial Sequence synthetic primer 20acggcttttg tatccaagtc ttttg 252120DNAArtificial SequenceDescription of Artificial Sequence synthetic primer 21gccctgtgcc ctccgctccc 202225DNAArtificial SequenceDescription of Artificial Sequence synthetic primer 22ggctcttcac atctctctct gcttc 252324DNAArtificial SequenceDescription of Artificial Sequence synthetic primer 23ccacactgac acctgctccc tttg 242422DNAArtificial SequenceDescription of Artificial Sequence synthetic primer 24cccgcaggtc caggctcagc tg 222523DNAArtificial SequenceDescription of Artificial Sequence synthetic primer 25cgcctccgcc gttaccgtcc ttg 232621DNAArtificial SequenceDescription of Artificial Sequence synthetic primer 26ccctggagcc ggcgcgcaaa g 212718DNAArtificial SequenceDescription of Artificial Sequence synthetic primer 27ccccgccccc gcccccgg 182821DNAArtificial SequenceDescription of Artificial Sequence synthetic primer 28cttcaggtcc ccccagtccc g 212926DNAArtificial SequenceDescription of Artificial Sequence synthetic primer 29ctagggatct tcagaggaag aaaaag 263025DNAArtificial SequenceDescription of Artificial Sequence synthetic primer 30gctgcgcggc gggtcagagc cccag 253141DNAArtificial SequenceDescription of Artificial Sequence synthetic primer 31ggccacgcgt cgactagtac tttttttttt tttttttttt n 413222DNAArtificial SequenceDescription of Artificial Sequence synthetic primer 32gaaaggcatc cgtaaggctc cc 223324DNAArtificial SequenceDescription of Artificial Sequence synthetic primer 33gacgccttta tgcatctgat tctc 243422DNAArtificial SequenceDescription of Artificial Sequence synthetic primer 34gaatcagatg cataaaggcg tc 223522DNAArtificial SequenceDescription of Artificial Sequence synthetic primer 35gggagcctta cggatgcctt tc 223620DNAArtificial SequenceDescription of Artificial Sequence synthetic primer 36agccccggct gctcgccagc 203724DNAArtificial SequenceDescription of Artificial Sequence synthetic primer 37ctgcgcggcg ggtcagagcc ccag 243820DNAArtificial SequenceDescription of Artificial Sequence synthetic primer 38agccccggct gctcgccagc 203924DNAArtificial SequenceDescription of Artificial Sequence synthetic primer 39gcctcagcag caaagcaaga tccc 244025DNAArtificial SequenceDescription of Artificial Sequence synthetic primer 40gctgagcctg gacctgttgg aaagg 254125DNAArtificial SequenceDescription of Artificial Sequence synthetic primer 41gctgagcctg gacctgttgg aaagg 254215PRTArtificial SequenceDescription of Artificial Sequence Synthetic peptide 42Cys Ser Lys Ser Phe Asp Gln Lys Ser Lys Asp Gly Asn Gly Gly 1 5 10 15431541DNAHomo sapiensCDS(43)..(612) 43gtgtccccgg agctgaaaga tcgcaaagag gatgcgaaag gg atg gag gac gaa 54 Met Glu Asp Glu 1ggc cag acc aaa atc aag cag agg cga agt cgg acc aat ttc acc ctg 102Gly Gln Thr Lys Ile Lys Gln Arg Arg Ser Arg Thr Asn Phe Thr Leu 5 10 15 20gaa caa ctc aat gag ctg gag agg ctt ttt gac gag acc cac tat ccc 150Glu Gln Leu Asn Glu Leu Glu Arg Leu Phe Asp Glu Thr His Tyr Pro 25 30 35gac gcc ttc atg cga gag gaa ctg agc cag cga ctg ggc ctg tcg gag 198Asp Ala Phe Met Arg Glu Glu Leu Ser Gln Arg Leu Gly Leu Ser Glu 40 45 50gcc cga gtg cag gtt tgg ttt caa aat cga aga gct aaa tgt aga aaa 246Ala Arg Val Gln Val Trp Phe Gln Asn Arg Arg Ala Lys Cys Arg Lys 55 60 65caa gaa aat caa ctc cat aaa ggt gtt ctc ata ggg gcc gcc agc cag 294Gln Glu Asn Gln Leu His Lys Gly Val Leu Ile Gly Ala Ala Ser Gln 70 75 80ttt gaa gct tgt aga gtc gca cct tat gtc aac gta ggt gct tta agg 342Phe Glu Ala Cys Arg Val Ala Pro Tyr Val Asn Val Gly Ala Leu Arg 85 90 95 100atg cca ttt cag cag gtt cag gcg cag ctg cag ctg gac agc gct gtg 390Met Pro Phe Gln Gln Val Gln Ala Gln Leu Gln Leu Asp Ser Ala Val 105 110 115gcg cac gcg cac cac cac ctg cat ccg cac ctg gcc gcg cac gcg ccc 438Ala His Ala His His His Leu His Pro His Leu Ala Ala His Ala Pro 120 125 130tac atg atg ttc cca gca ccg ccc ttc gga ctg ccg ctc gcc acg ctg 486Tyr Met Met Phe Pro Ala Pro Pro Phe Gly Leu Pro Leu Ala Thr Leu 135 140 145gcc gcg gat tcg gct tcc gcc gcc tcg gta gtg gcg gcc gca gca gcc 534Ala Ala Asp Ser Ala Ser Ala Ala Ser Val Val Ala Ala Ala Ala Ala 150 155 160gcc aag acc acc agc aag gac tcc agc atc gcc gat ctc aga ctg aaa 582Ala Lys Thr Thr Ser Lys Asp Ser Ser Ile Ala Asp Leu Arg Leu Lys165 170 175 180gcc aaa aag cac gcc gca gcc ctg ggt ctg tgacgccaac gccagcacca 632Ala Lys Lys His Ala Ala Ala Leu Gly Leu 185 190atgtcgcgcc tgtcccgcgg cactcagcct gcacgccctc cgcgccccgc tgcttctccg 692ttaccccttt gagacctcgg gagccggccc tcttcccgcc tcactgacca tccctcgtcc 752cctatcgcat cttggactcg gaaagccaga ctccacgcag gaccagggat ctcacgaggc 812acgcaggctc cgtggctcct gcccgttttc ctactcgagg gcctagaatt gggttttgta 872ggagcgggtt tgggggagtc tggagagaga ctggacaggg tagtgctgga accgcggagt 932ttggctcacc gcaaagctac aacgatggac tcttgcatag aaaaaaaaaa tcttgttaac 992aatgaaaaaa tgagcaaaca aaaaaatcga aagacaaacg ggagagaaaa agaggaaggc 1052aacttatttc ttaactgcta tttggcagaa gctgaaattg gagaaccaag gagcaaaaac 1112aaattttaaa attaaagtat tttatacatt taaaaatatg gaaaaacaac ccagacgatt 1172ctcgagagac tggggggagt taccaactta aatgtgtgtt ttaaaaaatg cgctaagaag 1232gcaaagcaga aagaagaggt atacttattt aaaaaactaa gatgaaaaaa gtgcgcaggt 1292gggaagttca caggttttga aactgacctt tttctgcgaa gttcacgtta atacgagaaa 1352tttgatgaga gaggcgggcc tccttttacg ttgaatcaga tgctttgagt ttaaacccac 1412catgtatgga agagcaagaa aagagaaaat attaaaacga ggagagagaa aaataatggc 1472aaaactgtct ggactgctga cagtaaattc cggtttgcat ggaaaaaaaa aaaaaaaaaa 1532aaaaaaaaa 15414410DNAHomo sapiens 44gtgatccacc 104520DNAHomo sapiens 45ccccacgcag ggatttatga 204620DNAHomo sapiens 46tctccccaag gtttggttcc 204720DNAHomo sapiens 47ttggacacag gcgtcatctt 204820DNAHomo sapiens 48gctcccgcag gtccaggctc 204920DNAHomo sapiens 49ttttttttag atggagtttt 205020DNAHomo sapiens 50gtggcagaag gtaagttcct 205120DNAHomo sapiens 51gcgcgtgcag gtaggaaccc 205220DNAHomo sapiens 52atgcataaag gtgggtgtcg 205320DNAHomo sapiens 53tttccaacag gtagctcact 205410DNAHomo sapiens 54aaaaaatagc 105510DNAHomo sapiens 55tgttacgtgg 10

Patent applications by Gudrun Rappold-Hoerbrand, Heidelberg DE

Patent applications in class 25 or more peptide repeating units in known peptide chain structure

Patent applications in all subclasses 25 or more peptide repeating units in known peptide chain structure

User Contributions:

Comment about this patent or add new information about this topic:

Inventors list

Assignees list

Classification tree browser

Top 100 Inventors

Top 100 Assignees

Patent application title: Human Growth Gene and Short Stature Gene Region

Inventors: Gudrun Rappold-Hoerbrand (Heidelberg, DE) Ercole Rao (Riedstadt, DE)
IPC8 Class: AA61K3827FI
USPC Class: 514 12
Class name: 25 or more peptide repeating units in known peptide chain structure
Publication date: 04/30/2009
Patent application number: 20090111744

Abstract:

Claims:

Description:

Inventors list

Assignees list

Classification tree browser

Top 100 Inventors

Top 100 Assignees

Patent application title: Human Growth Gene and Short Stature Gene Region

Inventors: Gudrun Rappold-Hoerbrand (Heidelberg, DE) Ercole Rao (Riedstadt, DE) IPC8 Class: AA61K3827FI USPC Class: 514 12 Class name: 25 or more peptide repeating units in known peptide chain structure Publication date: 04/30/2009 Patent application number: 20090111744

Abstract:

Claims:

Description:

Inventors: Gudrun Rappold-Hoerbrand (Heidelberg, DE) Ercole Rao (Riedstadt, DE)
IPC8 Class: AA61K3827FI
USPC Class: 514 12
Class name: 25 or more peptide repeating units in known peptide chain structure
Publication date: 04/30/2009
Patent application number: 20090111744