| Project/Area Number |
05454160
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| Research Category |
Grant-in-Aid for General Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Research Field |
General medical chemistry
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| Research Institution | Yamaguchi University |
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Principal Investigator |
NAKAZAWA Atsushi Yamaguchi University School of Medicine, Professor, 医学部, 教授 (90025594)
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| Co-Investigator(Kenkyū-buntansha) |
GOMADA Manabu Yamaguchi University School of Medicine, Research Assosiate, 医学部, 助手 (80243632)
INOUYE Sachiye Yamaguchi University School of Medicine, Assistant Professor, 医学部, 講師 (60159978)
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| Project Period (FY) |
1993 – 1994
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| Project Status |
Completed (Fiscal Year 1994)
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| Budget Amount *help |
¥5,900,000 (Direct Cost: ¥5,900,000)
Fiscal Year 1994: ¥1,900,000 (Direct Cost: ¥1,900,000)
Fiscal Year 1993: ¥4,000,000 (Direct Cost: ¥4,000,000)
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| Keywords | adenylate kinase / adenine nucleotide / gene targeting / isozyme / HeLa cell / ジーンターゲッティング / アイリザイム / 細胞内局在 |
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| Research Abstract |
Adenylate kinase is a ubiquitous enzyme which is involved in the interconversion of adenine nucleotides in the cells. In vertibrates, three isozymes (AK1, AK2, AK3) have been characterized. To understand the phisiological function of these isozyme, we have planned to isolate cell lines in which one or two AK genes were disrupted.
We showed HeLa cells have three functional AK genes by measuring enzyme activities, enzyme protein amounts, and mRNA levels.
Two targeting vectors for human AK1 gene were constructed. Plasmid AK1-Bsr contains a blasticidin S resistant gene and a diphtheria toxin A fragment gene. Plasmid AK-Neo contains a neomycin resistant gene and a diphtheria toxin A fragment gene. Using these plasmid, we shall be able to isolate AK1-disrupted HeLa cells by homologouse recombination.
To make a targeting vector for the human AK2 gene, we cloned cDNA of AK2 from HeLa cells using bovine AK2 cDNA as a plobe. Then, we cloned a part of gDNA of AK2, which will be useful to construct a gene targeting vector.
Isolation of cell lines which have disrupted AK genes are under investigation.
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