1990 Fiscal Year Final Research Report Summary
Introduction and expression of foreign gene in fish
| Project/Area Number |
01480080
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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 fisheries
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| Research Institution | Faculty of Agriculture, Miyazaki University |
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Principal Investigator |
AOKI Takashi Faculty of Agriculture, Miyazaki University, Professor, 農学部, 教授 (00051805)
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| Co-Investigator(Kenkyū-buntansha) |
OSHIRO Takashi Tokyo University of Fisheries, Associate Professor, 助教授 (10201401)
KITAO Tadatoshi Faculty of Agriculture Miyazaki University, Professor, 農学部, 教授 (60094075)
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| Project Period (FY) |
1989 – 1990
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| Keywords | fish globin gene / transgenic fish / expression of foreign gene in fish / promoter region / chloramphenicol acetyltransferase gene / 5'-flanking and 5'-untranslated regions / fish cell line / transfection |
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| Research Abstract |
The alpha-like globin genes from yellowtail (Seriola quinqueradiata) and carp (Cyprinus carpio) were cloned. The promoter elements of the -100 region, CCAAT box, and TATA box were found in the 5' -flanking region of fish globin genes. The cap site is located 46 base pairs upstream from ATG initiationcodon of the alpha-like globin gene of yellowtail and is 48 nucleotides upstream from the ATG sequences of the alpha-like globin gene of carp. The 5'-flanking and 5'-untranslated regions of yellowtail were ligated with chloramphenicol acetyltransferase (CAT) gene in vector plasmid. The recombinant plasmids were introduced into three kinds of fish tissue culture lines. The significantly CAT activity was detected in the fish tissue culture cell lines transfected with the 5'-flanking and 5'-untranslated regions-CAT recombinant plasmids. This result suggests the 5'-flanking region and 5'-untranslated regions from fish globin gene possess the promoter activity and play an important role in regulation of gene expression in fish tissue culture cells.
Carp alpha-globin gene (CalphaG) was microinjected into the cytoplasm of fertilized rainbow trout eggs at 1 cell stage treated with glutathione. About 67% of microinjected eggs hatched out and grew normally. CalphaG gene was detected from 130-day-old fish and 10 to 12-month-old fish. CalphaG-integrated and non-integrated cells coexisted mosaically in host fish. The CalphaG gene introduced into rainbow trout (Oncorhynchus mykiss) could be transmitted to their offspring.
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