Analysis of plant proteinase inhibitor gene and its use.
| Project/Area Number |
63560075
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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 |
応用生物化学・栄養化学
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| Research Institution | Hokkaido University |
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Principal Investigator |
OHNO Takeshi Hokkaido University, Faculty of Agriculture, Professor., 農学部, 教授 (00011726)
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| Project Period (FY) |
1988 – 1989
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| Project Status |
Completed (Fiscal Year 1989)
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| Budget Amount *help |
¥2,100,000 (Direct Cost: ¥2,100,000)
Fiscal Year 1989: ¥500,000 (Direct Cost: ¥500,000)
Fiscal Year 1988: ¥1,600,000 (Direct Cost: ¥1,600,000)
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| Keywords | Kunitz-type proteinase inhibitor / Chymotrypsin inhibitor / cDNA / Gene family / Signal peptide / Gene structure / Winged bean / 35S Promoter / クニッツ型キモトリプシンインヒビタ- / 35sプロモ-タ- / クニッツ型プロティナーゼインヒビター / cDNA塩基配列 / ゲノムライブラリー / プロモーター |
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| Research Abstract |
The nearly full-length cDNA to the mRNA encoding the major chymotrypsin inhibitor WCI-3 in winged bean seeds was cloned. It has an ORF composed of 207 amino acids, and the deduced sequence of the 183 C-terminal amino acids coincides precisely with the sequence determined for purified WCI-3.
Northern analysis revealed that the mRNA hybridizable to the WCI-3 cDNA started to accumulate in immature seeds 30 day after flowering. In addition, the hybridizable mRNA was detected a lot in tuberous roots and its cortices, a little in stems but not detected in shoots, leaves, root hairs and pods.
The chymotrypsin inhibitors were found to be encoded by a gene family, which is composed of at least 7 related genes per haploid, from the results of Southern analysis and the estimation of the diploid genome size of winged bean somatic cells. Until now 4 genes(WBG-1 - WBG-4) were isolated and analyzed. WBG-1 was supposed to be a pseudogene not coding for a functional inhibitor. WBG-3 had the sequence of w
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hich the 5' 2/3 is highly homologous to the cDNA but the remaining 1/3 completely different and the function was not known. WBG-2 and -4 are thought to code for functional chymotrypsin inhibitors different from WCI-3. WBG-4 encodes the protein identical to the WCI-3 except for one amino acid and is thought to be the gene encoding WCI-2 inhibitor protein. WBG-2 and -4, both are thought to be expressed, have the 5' upstream regions fairly homologous each other ranging over -1000 bp and it is interesting from the view point of regulation of gene expression.
The modified cauliflower mosaic virus 35S promoter, which is composed of tandemly repeated regions containing the enhancer element and tandemly doubled DNAs including TATA box, was shown to have the promoter activity much higher than the normal 35S promoter. The WCI-3 cDNA under the modified 35S promoter was introduced into tobacco plants using Ti vector system. The expression and the activity as an endogenous insecticide are now in progress. Less
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Report
(3 results)
Research Products
(7 results)
