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Molecular analysis of the mechanisms for controlling tannin accumulation of pollination-constant nonastringent (PCNA)-type persimmons in Chinese and Japanese cultivars to clarify different origins between them.

对中国和日本品种授粉恒定非涩性（PCNA）型柿子的单宁积累控制机制进行分子分析，以阐明它们之间的不同起源。

基本信息

批准号：
16380025
负责人：
YONEMORI Keizo
金额：
$ 10.05万
依托单位：
Kyoto University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2004
资助国家：
日本
起止时间：
2004 至 2006
项目状态：
已结题

项目摘要

The analysis was performed to identify key genes for regulating tannin accumulation into fruits of pollination constant non-astringent (PCNA)-type of persimmon originated in Japan and China. For this purpose, we conducted the following three experiments:1)We constructed genomic library from closely related diploid species, D. lotus, and a seed clone was screened from the library with a probe tightly linked to the trait of natural astringency-loss of Japanese-type PCNA fruits. Then, we constructed ca. 300 kbp contig of D. lotus. Due to the sequence analysis of this contig, we found that the sequences are very homologous to those of D. kaki and it can use for exploring the gene of persimmon (D. kaki). In addition, primer sets designed from the sequences of this contig were able to be applicable to distinguish Japanese-type PCNA from others. We could make a universal primer set to select PCNA persimmon.2)We conducted the expression analysis of mRNA to identify the genes involved in tannin accumulation of persimmon. For this purpose, suppression subtractive hybridization (SSH) analysis was performed between astringency-removed fruits with ethanol treatment on the tree and non-treated fruits. According to this analysis, we were able to obtain several candidates for the genes involved in tannin accumulation into the fruits.3)As for investigation of a dominant gene conferring the trait of Chinese-type PCNA, we looked for molecular markers by AFLP method combined with bulked segregant analysis (BSA) using progenies derived from a Chinese PCNA 'Luo Tian Tian Shi'. By this analysis, we found three markers closely linked to the Chinese-PCNA trait. Using a marker among them, we screened genomic library of `Luo Tian Tian Shi' and obtained a genomic clone as the seed for constructing contig for the survey of the gene.

本研究旨在对原产于日本和中国的授粉恒定非涩型柿果实单宁积累的调控基因进行分析。为此，我们进行了以下三个方面的实验：1）构建了近缘二倍体种D.莲的种子克隆，并从文库中筛选与日本型PCNA果实的天然涩味丧失性状紧密连锁的探针。然后，我们构建了CA。300 kbp的D.莲花通过对该重叠群的序列分析，我们发现该重叠群与D.可用于柿（D. kaki）。此外，从该重叠群的序列设计的引物组能够适用于区分日本型PCNA。2）通过mRNA的表达分析，鉴定了影响柿单宁积累的相关基因。为此目的，抑制消减杂交（SSH）分析之间的涩味去除果实与乙醇处理的树和未处理的水果。3）在研究中国型PCNA性状的显性基因方面，以中国型PCNA‘罗田田氏’的后代为材料，采用AFLP和BSA相结合的方法进行分子标记的研究。通过这种分析，我们发现了三个标记密切相关的中国PCNA性状。利用其中的一个标记，对‘罗田田氏’的基因组文库进行筛选，获得了一个基因组克隆，作为构建重叠群的种子，用于该基因的研究。