JP2002272467A - Base sequence of human kgf-like gene - Google Patents

Abstract

PROBLEM TO BE SOLVED: To determine the whole structure of a KGF-like gene transcription product participating in cancers and habitual abortion. SOLUTION: Various germs are each specified as an mRNA encoding a KGF-like protein and have each a determined base sequence.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

TECHNICAL FIELD The present invention relates to a novel human KGF-like gene involved in cancer and recurrent miscarriage and its nucleotide sequence. These genes are not only useful for diagnosis of cancer and recurrent miscarriage, but also provide a means for preventing or treating cancer and recurrent miscarriage.

[0002]

2. Description of the Related Art As a result of focusing attention on chromosome 9 inversion in ovarian cancer patients with a history of recurrent miscarriage, we have identified KGF-like genes as candidate genes for these diseases (Japanese Patent Application No. 11-028705, Yasuhara et al.). Although it was suggested that the chromosome 9 inversion of this patient might have occurred in the second and third exons of the KGF-like gene, the nucleotide sequence near the breakpoint was examined over about 50 kb on a normal gene, Only the sequence corresponding to exon 2 whose existence was already reported was recognized, and the presence of a nucleotide sequence showing a structure corresponding to exon 1 was not confirmed. That is, it has been suggested that this gene product similar to KGF may be involved in the occurrence and progression of cancer and miscarriage, but its structure and function remain unknown.

[0003]

As described above, elucidating the structure of the KGF-like gene and the function of its product is an effective means for clarifying the relationship between the above-mentioned disease and the KGF-like gene. Conceivable. It is also expected that the pathogenesis of these diseases will be analyzed from the obtained information to develop diagnosis and treatment of the diseases. However, only a part of the KGF-like gene structure has been reported so far (Proc. Natl. Ac
ad.Sci.USA, 89, 9287-9291 (1992), MJ Kelley
Et al.) Therefore, an object of the present invention is to determine the entire structure of a transcript of a KGF-like gene involved in cancer or habitual abortion.

[0004]

The start codon of the KGF gene is known to be present in exon 1 (Scie
nce, 245, 752-755 (1989), PW Finch et al.) For the KGF-like gene, only exons 2 and 3 have been reported, and the structure of the first exon was unknown (Proc. Natl. Acad. Sc
i.USA, 89, 9287-9291 (1992), MJ Kelley et al.). In addition, the KGF present in the inversion of chromosome 9
Exon 1 was absent in the same gene.
Therefore, in order to solve the above-mentioned problem, the 5′-terminal nucleotide sequence of KGF-like mRNA was determined using 5′-RACE or PCR. Accordingly, the present invention relates to various genes whose nucleotide sequences have been determined and identified as mRNAs encoding KGF-like proteins.

[0005]

BEST MODE FOR CARRYING OUT THE INVENTION The cDNA of the present invention can be obtained by the method described in the following examples. The gene of the present invention can be isolated from, for example, an existing human cDNA library using the cDNAs of SEQ ID NOS: 9 to 25 as probes. Alternatively, cDNA primers of SEQ ID NOS: 9 to 25 can be designed and isolated from human DNA or DNA libraries by PCR. The cDNA of the present invention is not limited to the gene sequences of SEQ ID NOS: 9 to 25. For example, the cDNA may be a gene having a certain modification or a gene having a certain homology with the above specific base sequence. Can be. The degree of the modification is not particularly limited, provided that the substance has the same function as the cDNA of SEQ ID NOS: 9 to 25. For example, of SEQ ID NO: 9 to 25
By modifying a part of cDNA by site-directed mutagenesis or the like, deletion, substitution, and addition derivatives can be freely produced.

[0006] The cDNA of the present invention can be used for diagnosis and treatment of cancer and recurrent miscarriage by checking the expression level of the KGF-like gene in various cells using the entire base sequence or a partial sequence thereof as a probe. In addition, primers are designed from the entire base sequence or a partial sequence thereof, and the expression level of the KGF-like gene in various cells is determined by PCR, whereby the primers can be used for diagnosis and treatment of cancer and repeated miscarriage. Also, using primer,
By analyzing mutations and deletions of the KGF-like gene, it can be used for diagnosis and treatment of cancer and recurrent miscarriage.

The cDNA of the present invention can express a sense or antisense gene by administering the whole nucleotide sequence or a partial sequence thereof to various cells, and can amplify or suppress the expression of a KGF-like gene. . Thereby, canceration and miscarriage can be controlled, and the therapeutic effect of an anticancer drug can be enhanced or maintained. In addition, a KGF-like protein can be obtained from this cDNA and its variant by a known method. The cDNA and the variant are expressively inserted into a suitable plasmid vector, virus vector, and vector such as a liposome vector, and a suitable host cell, for example, E. coli, yeast, insect cell, COS cell,
Mink lung epithelial cells, lymphocytes, fibroblasts, NIH / 3T3
Cells, CHO cells, blood cells, and tumor cells can be transformed, the host cells can be cultured, and a KGF-like protein or a derivative thereof can be produced from the culture. KG
The F-like protein or a partial peptide thereof can be used in the field of medicine as a drug containing this as an active ingredient. In addition, the KGF-like protein or partial peptide can be used as an immunogen for preparing an antibody, and an antibody against them can be freely prepared by a known method. This antibody can be used for diagnosis and treatment of cancer and miscarriage.

[0008]

EXAMPLES 1. Human KGF-like mRNA using 5'-RACE
5'-side nucleotide sequence determination Using Human Kidney Marathon-Ready cDNA (Clontech) as a template, a primer Ex3-R (SEQ ID NO: 1) having a sequence complementary to the nucleotide sequence near the stop codon in exon 3 of the KGF-like gene is gene-specific. PCR was performed using the primers. The other primer was AP-1 attached to cDNA, polymerase, buffer and dNTP Mix were manufactured by Clontech.
It was allowed to react at a temperature of less than or equal to shift conditions. That is, 94 ° C1
Hold at 94 ° C for 30 seconds, then hold at 72 ° C for 2 minutes 5 times, then at 94 ° C for 30 minutes.
After 70 seconds, 70 ° C. for 2 minutes was repeated 5 times, and finally, at 94 ° C. for 30 seconds and 68 ° C. for 2 minutes 25 times. The obtained PCR product was introduced into plasmid pCR2.1 using a TA cloning kit (Invitrogen).
E. coli Top 10 F 'strain was transformed with this plasmid
Plasmid DNA was purified from 144 strains, and after confirming the size of the inserted DNA fragment by EcoRI treatment, the nucleotide sequence was determined except for those in which the introduced DNA fragment was clearly small.

As a result, FIG. 1 and SEQ ID NOS: 9 to 25
The polymorphism of the KGF-like mRNA was confirmed as shown in FIG. That is,
The upstream region of Exon 2 existed with several units connected. The prototypes described on the right side of FIG. 1 are classified based on the polymorphisms of exon 2 and exon 3 (Pr
oc.Natl.Acad.Sci.USA, 89, 9287-9291 (1992), MJ K
elley et al.). When the amino acid sequence is predicted from mRNAs showing these polymorphisms, most of them are proteins such that the sequence encoded by the upstream exon is in frame with exons 2 and 3 due to the stop codon located immediately above exon 2. Will not be translated. Only clone K1-5
ATG that can be the start codon only in case of 6 (SEQ ID NO: 16)
There was no stop codon downstream of, and the sequence could be translated as a protein in frame with exons 2 and 3. Exon 2 near the inversion breakpoint reported earlier is a nucleotide sequence characterized by C at position 30 and G at position 32 from the 5 'end, and was considered not to belong to any prototype. (Figure 2). However, in all the clones whose nucleotide sequences were determined this time, the 32nd was all G, which is completely different from the 32nd being A in all reported prototypes. KGF near the inversion break point
Except for the 32nd difference, the similar gene corresponds to the base sequence of prototype II.

Clone K1-4 (SEQ ID NO: 10), K1-23
(SEQ ID NO: 13), a sequence present immediately above exon 2 common to K1-61 (SEQ ID NO: 17), K1-85 (SEQ ID NO: 20),
Approximately 100 bp was confirmed to be present at a position about 12 kb upstream of exon 2 by comparison with a base sequence of about 50 kb near the inversion cleavage point previously determined. On the other hand, when the base sequences of these clones were subjected to homology search, clones K1-1 (SEQ ID NO: 9), K1-10 (SEQ ID NO: 11), and K1-49 (SEQ ID NO: 1)
4) The region consisting of two units located immediately above exon 2 common to K1-142 (SEQ ID NO: 25) is CpG islan
The sequence was found to match the nucleotide sequence registered as d (NatureGenet. 6, 236-244 (1994), SH Cross
Et al.) Other than this sequence, no homology was recognized with a known sequence registered in the data bank.

2. Confirmation of KGF-like gene transcription by PCR In order to examine the expression status of mRNA polymorphism whose existence was suggested from the above-mentioned experiment, PCR was newly performed using a human cDNA panel (Clontech) as a template. PCR for confirming the length of the actually expressed mRNA was performed using the control cDNA of the panel as a template, and the primers Ex3-R (SEQ ID NO: 1) and KGF
Using a forward cDNA PCR kit (Clontech), using forward primers 1 to 7 (SEQ ID NOs: 2 to 8) designed from each unit of the same mRNA polymorphism.

The PCR reaction is maintained at 94 ° C. for 2 minutes and denatured at 94 ° C. for 40 seconds.
A cycle of annealing at 60 ° C. for 40 seconds and extending at 72 ° C. for 90 seconds was repeated 35 times. On the other hand, each cDNA for PCR The panel confirm the expression specificity in each organ as a template, the forward primer 1 (SEQ ID NO: 2) and Ex3-R (SEQ ID NO: 1)
Using forward of cDNA PCR kit as primers. K
The results of PCR on a human cDNA library using the base sequence of the GF-like gene are as shown in FIG. When a cDNA mixture prepared from each human organ is used as a template, DNA fragments of the expected lengths are amplified by forward primers 1 to 7 (SEQ ID NOs: 2 to 8) and Ex3-R (SEQ ID NO: 1), respectively. It was suggested that the result of the previous 5'-RACE was not an artifact. With respect to the amplification product of forward primer 1 (SEQ ID NO: 2) and Ex3-R (SEQ ID NO: 1), the base sequence was determined for several clones after subcloning into pCR2.1. Was confirmed. When cDNA derived from human organs was used as a template, PCR was performed using the forward primer 1 (SEQ ID NO: 2) and the Ex3-R (SEQ ID NO: 1) primer. As shown in the lower part of FIG. It was confirmed that various types of polymorphisms were universally expressed.

3. KGF-like for monoclomosomal cells
Genomic PCR of genes QIA amp tis from the human monochromosome cell line (Japanese Patent Application No. 11-028705, Yasuhara et al.) Used in the report of chromosome 9 inversion
Purify chromosomal DNA using sue kit (QIAGEN)
PCR was performed using age GC genomic PCR kit (Clontech). For PCR, SEQ ID NO: 1 and SEQ ID NO: 7 were used as primers. PCR reaction conditions were 95 ° C for 1 minute, 94
A cycle of denaturation at 30 ° C. for 30 seconds, annealing at 55 ° C. for 30 seconds and extension at 68 ° C. for 12 minutes was repeated 35 times, and finally kept at 68 ° C. for 12 minutes.

[0014] The KGF-like gene is not only chromosome 9 but also
It is also known to be present on chromosomes 5, 18, and 21.
On which chromosome the clones obtained this time were examined by genomic PCR. Forward primer 6
(SEQ ID NO: 7) and Ex3-R (SEQ ID NO: 1) was carried out using the P
The results of CR are shown in FIG. Among the monoclomosomal cells having chromosomes 2, 9, 15, 18, and 21, amplification products were confirmed only when genomic DNA purified from cell line 2209 having chromosome 9 was used as a template.

[0015]

[Sequence List] SEQUENCE LISTING <110> Asahi Breweries, Ltd. <120> Basic sequences of human KGF-like gene <130> 2000-18P <160> 25 <170> PatentIn Ver. 2.0 <210> 1 <211> 30 <212> DNA <213> Homo sapiens <400> 1 aagttattgc cataggaaga aagtgggctg 30 <210> 2 <211> 31 <212> DNA <213> Homo sapiens <400> 2 tgacccagtc agtgcccttc agttaaagac c 31 <210> 3 <211> 28 <212> DNA <213> Homo sapiens <400> 3 cgcatagact ttacctgcgc acgcgtcg 28 <210> 4 <211> 33 <212> DNA <213> Homo sapiens <400> 4 ccctaaagga tgatctgtct tctcatcatt ttg 33 <210> 5 <211 > 29 <212> DNA <213> Homo sapiens <400> 5 gatgtaacaa aacaaaggag tcttcaatg 29 <210> 6 <211> 32 <212> DNA <213> Homo sapiens <400> 6 catgctcaag ctatcctctg accttagcct cc 32 <210> 7 < 211> 32 <212> DNA <213> Homo sapiens <400> 7 catctgtacg gttgggacaa gtacagtaac cc 32 <210> 8 <211> 28 <212> DNA <213> Homo sapiens <400> 8 cgcatagact ttacctgcgc acgcgtcg 28 <210> 9 <211> 790 <212> DNA <213> Homo sapiens <400> 9 gaaaacgggc aatgggtgga ttcgggtcca gattctggca ggagggagtt tgggatcga g 60 atctggaaaa aagcactaga ctggaagagg acgcgatgga gtcggagccg ctngcgggga 120 caaaaaccag aggccgggna aggcgccggt gggaggcaag gcgcgcatag actttacctg 180 cgcacgcgtc gcagccatct ccgtgcacag tggtggccac cgcgactggt gctgaagtgt 240 cggcgcgtgc cgggcgctcc gctgggaccc gggttgctcg ccctgagtct cagctttctc 300 atctgtacgg ttgggacaag tacagtaacc ctcgcccgtc aagacgggcc agggctgtgg 360 cgagggtcca cgccttagag caggcaccta tcttgtgcag ggccctgaga cggggtctga 420 ctcagttcct gcggggaact tcaccagtga cccagtcagt gcccttcagt taaagaccac 480 caggagcaca cttatatcat ggaaatcagg acagtggcag ttgggattgt ggcaatcaaa 540 ggggtggaaa gtgaattcta tcttgcaacg aacgaggaag gaaaactcta tgcagagaag 600 gaatacaatg aagattgtaa cttcaaagat ctaattctgg aaaaccatta caacacatat 660 gcagcagcta aatggacaaa caacggaggg gaaatgtttg tggccttaaa tcaaaagggg 720 attcctgtaa gaggaaaaaa aaaaaagaaa gaacaaaaaa cagcccactt tcttcctatg 780 gcaataactt 790 <210> 10 <211> 577 <212> DNA <213> Homo sapiens <400> 10 aaaaaaaaaa aaaaaaagcg accgctgcaa gacgggccag ggctgtggcg agggtccacg 60 ccttagagca ggc acctatc ttgtgcaggg ccctgagacg gggtctgact cagttcctgc 120 ggggaacttc accagtgacc cagtcagtgc ccttcagtta aagaccacca ggagcacact 180 tgctggagtg cagtggcatg accatggctc actacagtct tgaactttca tgctcaagct 240 atcctctgac cttagcctcc tgagtagctg gaactacaga tatcatggaa atcaggacag 300 tagcagttgg gattgtggca atcaaagggg tggaaagtga attctatctt gcaacgaacg 360 aggaaggaaa actctatgca aagaaggaat acaatgaaga ttgtaacttc aaagatctaa 420 ttctggaaaa ccattacaac acatatgcag cagctaaatg gacaancaac ggaggggaaa 480 tgtttgtggc cttaaatcaa aaggggattc ctgtaagagg aaaaaaaaaa aaagaaagaa 540 caaaanncag cccactttct tcctatggca ataactt 577 <210> 11 <211> 581 <212> DNA <213> Homo sapiens <400> 11 caccgcgact ggtgctgaag tgtcggcgtg tgccgggcgc tccgctggga cccgggttgc 60 tcgccctgag tctcagcttt ctcatctgta cggttgggac aagtacagta accctcgccc 120 gtcaagacgg gccagggctg tggcgagggt ccacgcctta gagcaggcac ctatcttgtg 180 cagggccctg agatggggtc tgactcagtt cctgcgggga acttcaccag tgacccagtc 240 agtgcccttc agttaaagac caccaggagc acacttatat catggaaatc aggacagtgg 300 cag ttcggat tgtggcaatc aaaggggtgg aaagtgaatt ctatcttgca atgaacgagg 360 aaggaaaact ctatgcaaag aaagaatgca atgaagattg taacttcaaa gaactaattc 420 tggaaaacca ttacaacaca tatgcagcag ctaaatggac acacaatgga ggggaaatgt 480 ttgttgcctt aaatcaaaag gggattcctg taagaggaaa aaaaacgaag aaagaacaaa 540 aaacagccca ctttcttcct atggcaataa cttaagccga a 581 <210> 12 <211> 683 <212> DNA <213> Homo sapiens <400> 12 caccgcgact ggtgctgaag tgtcggcgtg tgccgggcgc tccgctggga cccgggttgc 60 tcgccctgag tctcagcttt ctcatctgta cggttgggac aagtacagta accctcgccc 120 gtcaagacgg gccagggctg tggcgagggt ccacgcctta gagcaggcac ctatcttgtg 180 cagggccctg agatggggtc tgactcagtt cctgcgggga acttcaccag tgacccagtc 240 agtgcccttc agttaaagac caccaggagc acacttcatt gctgttggaa aggctgatgc 300 caatcaaaac caatttttcc ctaaaggatg atctgtcttc tcatcatttt gacaggatgt 360 aacaaaacaa aggagtcttc aatgttatat catggaaatc aggacagtgg cagttcggat 420 tgtggcaatc aaaggggtgg aaagtgaatt ctatcttgca atgaacgagg aaggaaaact 480 ctatgcaaag aaagaatgca atgaagattg taacttcaaa gaactaattc tg gaaaacca 540 ttacaacaca tatgcagcag ctaaatggac acacaatgga ggggaaatgt ttgttgcctt 600 aaatcaaaag gggattcctg taggaggaaa aaaaacgaag aaagaacaaa aaacagccca 660 ctttcttcct atggcaataa ctt 683 <210> 13 <211> 388 <212> DNA <213> Homo sapiens <400> 13 tgcagtggca tgaccatggc tcactacagt cttgaacttt catgctcaag ctatcctctg 60 actttagcct cctgagtagc tggaactaca gatatcatgg aaatcaggac agtagcagtt 120 gggattgtgg caatcaaagg ggtggaaagt gaattctatc ttgcaacgaa cgaggaagga 180 aaactctatg caaagaagga atgcaatgaa gattgtaact tcaaagatct aattctggaa 240 aaccattaca acacatatgc agcagctaaa tggacaaaca acggagggga aatgtttgtg 300 gccttaaatc aaaaggggat tcctgtaaga ggaaaaaaaa caaagaaaga acaaaaaaca 360 gcccactttc ttcctatggc aataactt 388 <210> 14 <211> 949 <212> DNA < 213> Homo sapiens <400> 14 cagtaacaaa catggctccc tgaagccgct caggctcaag agcaacatgg aggtctgcac 60 ttaatcgctc ctctccgggg gcggccatac tgaggaggca tctggtccgt gcagg aggtcagca tggcc acctcaag gg gat ctggaaaaaa gcactagact 240 ggaagaggac gcgatggagt cggagccgct ggcggggaca aaaaccagag gccggggaag 300 gcgccggtgg gaggcaaggc acggatggac tttacctgcg cacgcgtcgc agccatctcc 360 gcgcacagtg gtggcccacc gcgactggtg ctgaagtgtc ggcgcgtgcc gggcgctccg 420 ctgggacccg ggttgctggc cctgagtgtc agctttctca tctgtacggt tgggacaagt 480 acagtaaccc tcgcccgtca agacgggcca gggctgtggc gagggtccac gccttagagc 540 aggcacctat cttgtgcagg gccctgagat ggggtctgac tcagttcctg cggggaactt 600 caccagtgac ccagtcagtg cccttcagtt aaagaccacc aggagcacac ttatatcatg 660 gaaatcagga cagtagcagt tgggattgtg gcaatcaaag gggtggaaag tgaattctat 720 cttgcaacga acgaggaagg aaaactctat gcaaagaagg aatgcaatga agattgtaac 780 ttcaaagatc taattctgga aaaccattac aacacatatg cagcagctaa atggacaaac 840 aacggagggg aaatgtttgt ggccttaaat caaaagggga ttcctgtaag aggaaaaaaa 900 acaaagaaag aacaaaaaac agcccacttt cttcctatgg caataactt 949 <210> 15 <211> 729 <212> DNA < 213> Homo sapiens <400> 15 ggacccgggt tgctcgccct gagtctcagc tttctcatct gtacggttgg gacaagtaca 60 gtaaccctcg cccgt caaga cgggccaggg ctgtggcgag ggtccacgcc ttagagcagg 120 cacctatctt gtgcagggcc ctgagatggg gtctgactca gttcctgcgg ggaacttcac 180 cagtgaccca gtcagtgccc ttcagttaaa gaccaccagg agcacactta gatggggttt 240 cgccttgctg cccacgctgg tcctgaaccc ctgagctcag gtgatccacc cacctcggcc 300 tcccatactg ctgggattac agcattgctg ttggaaaggc tgatgccaat caaaaccaat 360 ttttccctaa aggatgatct gtcttctcat cattttgaca ggatgtaaca aaacaaagga 420 gtcttcaatg ttatatcatg gaaatcagga cagtagcagt tgggattgtg gcaatcaaag 480 gggtggaaag tgaattctat cttgcaacga acgaggaagg aaaactctat gcaaagaagg 540 aatgcaatga agattgtaac ttcaaagatc taattctgga aaaccattac aacacatatg 600 cagcagctaa atggacaaac aacggagggg aaatgtttgt ggccttaaat caaaagggga 660 ttcctgtaag aggaaaaaaa acaaagaaag aacaaaaaac agcccacttt cttcctatgg 720 caataactt 729 <210> 16 <211> 468 <212> DNA <213> Homo sapiens <400> 16 ctagaattca gcggccgttt cttagagcag gcacctatct tgtgcagggc cctgagatgg 60 ggtctgactc agttcctgcg gggaacttca ccagtgaccc agtcagtgcc cttcagttaa 120 agaccaccag gagcacactt gatgtaacaa aac aaaggag tcttcaatgt tatatcatgg 180 aaatcaggac agtggcagtt cggattgtgg caatcaaagg ggtggaaagt gaattctatc 240 ttgcaatgaa cgaggaagga aaactctatg caaagaaaga atgcaatgaa gattgtaact 300 tcaaagaact aattctggaa aaccattaca acacatatgc agcagctaaa tggacacaca 360 atggagggga aatgtttgtt gccttaaatc aaaaggggat tcctgtaaga ggaaaaaaaa 420 cgaagaaaga acaaaaaaca gcccactttc ttcctatggc aataactt 468 <210> 17 <211> 418 <212> DNA <213> Homo sapiens <400> 17 catgttttta ccctgttgcc caggctggag tgcagtggca tgaccatggc tcactacagt 60 cttgaacttt catgctcaag ctatcctctg accttagcct cctgagtagc tggaactaca 120 gatatcatgg aaatcaggac agtagcagtt gggattgtgg caatcaaagg ggtggaaagt 180 gaattctatc ttgcaacgaa cgaggaagga aaactctatg caaagaagga atacaatgaa 240 gattgtaact tcaaagatct aattctggaa aaccattaca acacatatgc agcagctaaa 300 tggacaaaca acggagggga aatgtttgtg gccttaaatc aaaaggggat tcctgtaaga 360 ggaaaaaaaa caaagaaaga acaaaaaaca gcccactttc ttcctatggc aataactt 418 <210> 18 <211> 779 <212> DNA <213> Homo sapiens <400> 18 ccgcgactgg tgctgaagtg tcggc gcgtg ccgggcgctc cgctgggacc cgggttgctc 60 gccctgagtc tcagctttct catctgtacg gttgggacaa gtacagtaac cctcgcccgt 120 caagacgggc cagggctgtg gcgagggtcc acgccttaga gcaggcacct atcttgtgca 180 gggccctgag acggggtctg actcagttcc tgcggggaac ttcaccagtg acccagtcag 240 tgcccttcag ttaaagacca ccaggagcac acttcattgc tgttggaaag gctgatgcca 300 atcaaaacca atttttccct aaaggatgat ctgtcttctc atcattttga caggatgtaa 360 caaaacaaag gagtcttcaa tgttgctgga gtgcagtggc atgaccatgg ctcactacag 420 tcttgaactt tcatgctcaa gctatcctct gaccttagcc tcctgagtag ctggaactac 480 agatatcatg gaaatcagga cagtagcagt tgggattgtg gcaatcaaag gggtggaaag 540 tgaattctat cttgcaacga acgaggaagg aaaactctat gcaaagaagg aatacaatga 600 agattgtaac ttcaaagatc taattctgga aaaccattac aacacatatg cagcagctaa 660 atggacaaac aacggagggg aaatgtttgt ggccttaaat caaaagggga ttcctgtaag 720 aggaaaaaaa acaaagaaag aacaaaaaac agcccacttt cttcctatgg caataactt 779 <210> 19 <211> 998 <212> DNA <213> Homo sapiens <400> 19 tgcctgttga tttatggaaa caattatgat tctgctggag aacttttcag ctgagaa ata 60 gtttgtagct acagtagaaa ggctcaagtt gcaccaggca gacaacagac atggaattct 120 tatatatcca gctgttagca acaaaacaaa agtcaaatag caaacagcgt cacagcaact 180 gaacttacta cgaactgttt ttatgaggat ttatcaacag agttatttaa ggaggaatcc 240 tgtgttgtta tcaggaacta aaaggataag gctaacaatt tggaaagagc aactactctt 300 tcttaaatca atctacaatt cacagatagg aagaggtcaa tgacctagga gtaacaatca 360 actcaagatt cattttcatt atgttattca tgaacacccg gagcactaca ctataatgca 420 caaatggata ctgacatgga tcctgccaac tttgctctac agatcatgct ttcacattat 480 ctgtctagtg ggtactatat ctttagcttg caatgacatg actccagagc aaatggctac 540 aaatgtgaac tgttccagcc ctgagcgaca cacaagaagt tatgattaca tggaaggagg 600 ggatataaga gtgagaagac tcttctgtcg aacacagtgg tacctgagga tcgataaaag 660 aggcaaagta aaagggaccc aagagatgaa gaataattac aatatcatgg aaatcgggac 720 agtggcagtt ggaattgtgg caatcaaagg ggtggaaagt gaattctatc ttgcaatgaa 780 caaggaagga aaactctatg caaagaaaga atgcaatgaa gattgtaact tcaaagaact 840 aattctggaa aaccattaca acacatatgc atcagctaaa tggacacaca acggagggga 900 aatgtttgtt gc cttaaatc aaaaggggat tcctgtaaga ggaaaaaaaa cgaagaaaga 960 acaaaaaaca gcccactttc ttcctatggc aataactt 998 <210> 20 <211> 669 <212> DNA <213> Homo sapiens <400> 20 ccgcgactgg tgctgaagtg tcggcgcgtg ccgggcgctc cgctgggacc cgggttgctc 60 gccctgagtc tcagctttct catctgtacg gttgggacaa gtacagtaac cctcgcccgt 120 caagacgggc cagggctgtg gcgagggtcc acgccttaga gcaggcacct atcttgtgca 180 gggccctgag acggggtctg actcagttcc tgcggggaac ttcaccagtg acccagtcag 240 tgcccttcag ttaaagacca ccaggagcac acttgctgga gtgcagtggc atgaccatgg 300 ctcactacag tcttgaactt tcatgctcaa gctatcctct gaccttagcc tcctgagtag 360 ctggaactac agatatcatg gaaatcagga cagtagcagt tgggattgtg gcaatcaaag 420 gggtggaaag tgaattctat cttgcaacga acgaggaagg aaaactctat gcaaagaagg 480 aatacaatga agattgtaac ttcaaagatc taattctgga aaaccattac aacacatatg 540 cagcagctaa atggacaaac aacggagggg aaatgtttgt ggccttaaat caaaagggga 600 ttcctgtaag aggaaaaaaa aaaaagaaag aacaaaaaac agcccacttt cttcctatgg 660 caataactt 669 <210> 21 <211> 467 <212> DNA <213> Homo sapiens <400> 21 aaaaaaaaaa ggcggccgct gaatcagtgc ccttcagtta aagaccacca ggagcacact 60 tcattgctgt tggaaaggct gatgccaatc aaaaccaatt tttccctaaa ggatgatctg 120 tcttctcatc attttgacag gatgtaacaa aacaaaggag tcttcaatgt tatatcatgg 180 aaatcaggac agtggcagtt cggattgtgg caatcaaagg ggtggaaagt gaattctatc 240 ttgcaatgaa cgaggaagga aaactctatg caaagaaaga atgcaatgaa gattgtaact 300 tcaaagaact aattctggaa aaccattaca acacatatgc agcagctaaa tggacacaca 360 atggagggga aatgtttgtt gccttaaatc aaaaggggat tcctgtaaga gnnaaaaaac 420 gaagaaagaa caaaaaacag cccactttct tcctatggca ataactt 467 <210> 22 <211> 640 <212> DNA <213> Homo sapiens <400> 22 ataatctcaa agtcattagc agaggtctat atttcagggt agctgtcaga agctttttat 60 ttttttcatg aaccttcttg aagaaaaaaa aaaaaaaaaa aaaaaaaaaa gcggcgccgc 120 tgaattctga acctgagtct cagctttctc atctgtacgg ttgggacaag tacagtaacc 180 ctcgcccgtc aagacgggcc agggctgtgg cgagggtcca cgccttagag caggcaccta 240 tcttgtgcag ggccctgaga tggggtctga cccagttcct gcggggaact tcaccagtga 300 cccagtcagt gcccttcagt taaagaccac caggagcaca ctta tatcat ggaaatcagg 360 acagtggcag ttcggattgt ggcaatcaaa ggggtggaaa gtgaattcta tcttgcaatg 420 aacgaggaag gaaaactcta tgcaaagaaa gaatgcaatg aagattgtaa cttcaaagaa 480 ctaattctgg aaaaccatta caacacatat gcagcagcta aatggacaca caatggaggg 540 gaaatgtttg ttgccttaaa tcaaaagggg attcctgtaa gaggaaaaaa aaacgaagaa 600 agaacaaaaa cagcccactt tcttcctatg gcaataactt 640 <210> 23 <211> 721 <212> DNA <213 > Homo sapiens <400> 23 ccatcctaat acgactcact atagggcttc gagcggccgc ccgggcaggt gacaattagt 60 aaatctgttt gaagtcaaag gatcttctaa gttgtagaag aaaaatggga gaaaattatg 120 taggctggag tcagattttg gagagctttg gatagtaaat aaaagaggtt tttgtttgct 180 tttgttttta atagagtata ccactgaaac ttttgaagca gaagagtaac ctttataaca 240 tatatcttat gaatacataa agggttaaag tcagaagggc tggaggcaaa ggtctagcta 300 agaaactata ataatagttg aggcacaaga aaatgaaggc ctggactaat gtgatggcaa 360 tggaaacaga agagaacggc tggatataca ttggtaatat agagaggaaa cataattgtt 420 tctgatatca tggaaatcag gacagtggca gttggaattg tggcaatcaa aggggtggaa 480 agtgaattct atcttgcaat gaacaaggaa g gaaaactct atgcaaagaa agaatgcaat 540 gaagattgta acttcaaaga actaattctg gaaaaccatt acaacacata tgcatcagct 600 aaatggacac acaacggagg ggaaatgttt gttgccttaa atcaaaaggg gattcctgta 660 agaggaaaaa aaacgaagaa agaacaaaaa acagcccact ttcttcctat ggcaataact 720 t 721 <210> 24 <211> 626 <212> DNA <213> Homo sapiens <400> 24 ccatcctaat acgactcact atagggctcg agcggccgcc cgggcaggtg taaccctcgc 60 ccgtcaagac gggccagggc tgtggcgagg gtccacgcct tagagcaggc acctatcttg 120 tgcagggccc tgagacgggg tctgactcag ttcctgcggg gaacttcacc agtgacccag 180 tcagtgccct tcagttaaag accaccagga gcacacttca ttgctgttgg aaaggctgat 240 gccaatcaaa accaattttt ccctaaagga tgatctgtct tctcatcatt ttgacaggat 300 gtaacaaaac aaaggagtct tcaatgttat atcatggaaa tcaggacagt agcagttggg 360 attgtggcaa tcaaaggggt ggaaagtgaa ttctatcttg caacgaacga ggaaggaaaa 420 ctctatgcaa agaaggaata caatgaagat tgtaacttca aagatctaat tctggaaaac 480 cattacaaca catatgcagc agctaaatgg acaaacaacg gaggggaaat gtttgtggcc 540 ttaaatcaaa aggggattcc tgtaagagga aaaaaaacan agaaagaaca aaaaaca gcc 600 cacttttctt cctatggcaa taactt 626 <210> 25 <211> 632 <212> DNA <213> Homo sapiens <400> 25 cctaatacga ctcactatag ggctcagcgn ccgcccgggc aggtgcgcac agtggtggcc 60 accgcgactg gtgctgaagt gtcggcgcgt gccgggcgct ccgctgggac ccgggttgct 120 ggccctgagt gtcagctttc tcatctgtac ggttgggaca agtacagtaa ccctcgcccg 180 tcaagacggg ccagggctgt ggcgagggtc cacgccttag agcaggcacc tatcttgtgc 240 agggccctga gatggggtct gactcagttc ctgcggggaa cttcaccagt gacccagtca 300 gtgcccttca gttaaagacc accaggagca cacttatatc atggaaatca ggacagtggc 360 agttcggatt gtggcaatca aaggggtgga aagtgaattc tatcttgcaa tgaacgagga 420 aggaaaactc tatgcaaaga aagaatgcaa tgaagattgt aacttcaaag aactaattct 480 ggaaaaccat tacaacacat atgcagcagc taaatggaca cacaatggag gggaaatgtt 540 tgttgcctta aatcaaaagg ggattcctgt aagaggaaaa aaaacgaaga aagaacaaaa 600 aacagcccac tttcttccta tggcaataac tt 632

[Brief description of the drawings]

FIG. 1 shows the classification of KGF-like cDNA clones. In the figure, boxes with the same pattern show almost the same base sequence. The numbers shown in the table on the right prototype indicate the number of clones obtained by the prototype based on the nucleotide sequence of exons 2 and 3, and structurally based on the upstream sequence. The prototype III 'will be described with reference to FIG. Clone KGF represented by K1-79.

FIG. 2 shows the nucleotide sequence of exon 2 of each prototype. inv: Nucleotide sequence of exon 2 of the KGF-like gene present in about 50 kb near the breakpoint of chromosome 9. I, II, III, I
V: Exon 2 base sequence of each prototype (3). Prototype I indicates KGF.

FIG. 3 shows the expression of KGF-like mRNA. Upper: DNA fragment amplified from human cDNA Mix with any of forward primers 1 to 7 and Ex3-R was subjected to agarose gel electrophoresis.
Lower: Forward primer 1 using cDNA from human organs as template
A DNA fragment amplified using Ex3 and Ex3-R as primers was subjected to electrophoresis on an agarose gel.

FIG. 4 is a diagram showing genomic PCR of a KGF-like gene for monoclomosomal cells. Lane 1-6: chromosomal DNA template was purified from each cell line, lane 7: template distilled water.

──────────────────────────────────────────────────続 き Continued on the front page (51) Int.Cl. ⁷ Identification symbol FI theme coat ゛ (Reference) C07K 14/47 C12N 1/15 C12N 1/15 1/19 1/19 1/21 1/21 15/00 ZNAA 5/10 5/00 A (72) Inventor Seiji Isonishi 3-5-7 Narita Nishi, Suginami-ku, Tokyo (72) Inventor Aiko Okamoto 4-5-2 Igusa, Suginami-ku, Tokyo F-term (reference) 4B024 AA01 AA11 BA80 CA04 CA20 DA02 DA06 DA12 GA11 HA11 HA17 4B065 AA01X AA26X AA57X AA58X AA72X AA88X AA90X AB01 AC14 BA02 CA24 CA44 CA46 4C084 AA13 NA14 ZA812 ZB262 4C086 AA01 AA20 A14A01 AA04 AA01 AA04 AA01 AA04 AA01 MAA

Claims (12)

[Claims] 1. A human KGF-like genes characterized in that it has a nucleotide sequence of SEQ ID NO: 9-25. 2. A cDN having the nucleotide sequence of SEQ ID NOS: 9 to 25.
A human KGF-like gene, wherein A expresses a hybridizing mRNA. 3. SEQ ID NOS: 9 to 25 involved in KGF-like action
The KGF-like gene shown in. 4. A KGF-like gene represented by SEQ ID NOS: 9 to 25 involved in cancer development. 5. The KGF-like gene shown in SEQ ID NOS: 9 to 25, which is involved in recurrent miscarriage. 6. A vector comprising the nucleotide sequence according to claim 1 or 2. 7. The vector according to claim 6, wherein the vector is selected from the group consisting of a plasmid vector, a virus vector, and a liposome vector. 8. A host cell transformed by the vector according to claim 6 or 7. 9. A cell selected from the group consisting of Escherichia coli, yeast, insect cells, COS cells, mink lung epithelial cells, lymph cells, fibroblasts, NIH / 3T3 cells, CHO cells, blood cells, and tumor cells. in it, the host cell of claim 8. 10. A gene therapy agent for malignant tumor comprising the nucleotide sequence according to claim 1 or 2. 11. An agent for recurrent miscarriage gene therapy comprising the nucleotide sequence according to claim 1 or 2. 12. A protein expressed from the gene according to claim 1.