1995 Fiscal Year Final Research Report Summary
Isolation of a new gene by a PCR-cloning method.
| Project/Area Number |
05671009
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| Research Category |
Grant-in-Aid for General Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Research Field |
General surgery
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| Research Institution | Ehime University |
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Principal Investigator |
KIMURA Shigeru School of Medicine, Ehime University.Professor, 医学部, 教授 (30036462)
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| Co-Investigator(Kenkyū-buntansha) |
WATANABE Yuji School of Medicine, Ehime University.Assistant, 医学部, 助手 (20210958)
SATO Motomichi School of Medicine, Ehime University.Lecture, 医学部, 講師 (50162491)
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| Project Period (FY) |
1993 – 1995
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| Keywords | PCR / differential display / cloning / cancer cell lines |
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| Research Abstract |
A lot of oncogenes and tumor suppressor genes were discovered in past several years, but these genes are not enough to explain the mechanism of carcinogenesis. There are a number of important genes that regulate cell proliferation and differentiation, and certain genetic changes of such genes cause a malignant transformation. Most of them have not been discovered yet. In this research, we tried to isolate a new gene concerning cell differentiation or proliferation.
We used a differential display technique to identify several cDNA sequences. This method is rapid to detect differences of expression of mRNA between one material and the other. We prepared cancer cell lines treated with or without PMA (Phorbol Myristate Acetate), isolated mRNA from each of them, and synthesized cDNA.Consequence of DNA sequencing, we obtained some cDNA fragments not recorded in the genebank. We examined their expression patterns using Northern blotting in several cancer cell lines, and tried to isolate the complete cDNA of these. Finally, we obtained three attracted cDNA fragments of unknown genes.
The clone9 which was isolated from LNCaP,derived from prostatic cancer cell line, was expressed strongly in all examined cell lines including leukemia, brest, colon, prostate and cervix of uterus. The size of mRNA is estimated about 1.7kb, and its amino acid sequence demonstrated partial homology with tyrosine-phosphatase.
The clone16-5 was also isolated from LNCaP which demonstrated remarkable expression broadly in our cell lines. The size of mRNA is estimated about 3.7kb.
The clone15-1 showed interesting expression pattern. Its expression was only observed in LNCaP treated with PMA,the size of mRNA was predicted 3.8kb.This gene may correlate with a cell differentiation or proliferation.
We will try to isolate complete cDNA of these genes, and characterize of their functions.
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[Publications] Kito, K., Kihara, T., Ueda, N., Aono, K., Miyauchi, K., Kimura, S.and Hashimoto, K.: "Absence of K-ras, Ha-ras and p53 gene mutations in neuroblastoma.Jpn." J.Pediatr.Oncol.29. 295-297 (1992)
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