1997 Fiscal Year Final Research Report Summary
Molecular analysis of self-incompatibility gene product in tea-plants.
| Project/Area Number |
08660007
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Breeding science
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| Research Institution | Mie University |
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Principal Investigator |
KOWYAMA Yasuo Mie University, Faculty of Bioresources, Professor, 生物資源学部, 教授 (80024579)
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| Co-Investigator(Kenkyū-buntansha) |
KAKEDA Katsuyuki Mie University, Faculty of Bioresources, Associate Professor, 生物資源学部, 助教授 (50221867)
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| Project Period (FY) |
1996 – 1997
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| Keywords | Tea-plant / Pistil-specific protein / Pathogenesis-related / RNA binding protein / Self-incompatibility |
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| Research Abstract |
Tea-plants show a gametophytic type of self-incompatibility that is controlled by a single locus with multiple alleles (called S-gene). The present study is undertaken to identify some proteins specific for pistil tissue of tea-plants, and to elucidate a molecular mechanism underlysing in the self-incompatibility. From analysis of pistil proteins after 2D-PAGE,we detected two major proteins, A (ca.18kDa) and B (ca.15kDa) which showed a polymorphic pattern in pH and molecular weigh among tea cultivars different in S-genotypes. Following PVDF-mambrane transfer of the 2D-PAGE gel, N-terminal amino acid sequence of protein A spot was determined by a protein sequencer. Based on the N-terminal amino acid sequence, degenerated oligonucleotide-primers were synthesized. Using these primers, RT-PCR (3'-RACE) was performed with pistil mRNA.PCR product was confirmed to have the N-terminus corresponding nucleotide sequence, and used as a probe for screening of pistil cDNA library. A full-length cDNA clone obtained was found to encode pathogenesis related protein (PR-1) that showed 40% amino acid identity with tobacco PR-1. Northern blot analysis showed that gene expression of the PR-1 was specific for pistil tissue with no detection in stamen and leaf. N-terminus of Protein B was also sequenced, but it's N-terminus was blocked. So, we determined some internal amino acid sequences of peptides obtained from HPLS-fractionation after lysil-endopeptidase digestion of the protein B spot. From homology search of these amino acid sequences in the Gen Bank data base, protein B was found to be a member of glycine-rich RNA binding proteins that are known to be induced by stress conditions such as cold temperature in many plant species. Biological functions of the two pistil proteins indentified in this study are not known in tea-plants. Further study is needed to make clear whether these proteins are involved in the self-incompatibility reaction of tea-plants.
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