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Isolation and transfer of genes of key enzymes to improve the quality of fruit

提高果实品质关键酶基因分离与转移

基本信息

批准号：
06304013
负责人：
YAMAKI Shohei
金额：
$ 14.27万
依托单位：
Nagoya University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (A)
财政年份：
1994
资助国家：
日本
起止时间：
1994 至 1996
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-06304013/
关键词：
ACCsynthase cDNA Cysteine protease gene Sucrose phosphate synthase cDNA Sorbitol dehydrogenase cDNA S6PDH transgenic apple Transgenic grape Transgenic citrus Transgenic persimmon ACC合成酵素遺伝子 ACC酸化酵素遺伝子 S6PDH遺伝子 SPS遺伝子リンゴの形質転換体ソル

项目摘要

This project was conducted by 2 groups. The first group tried the purification and cloning of some enzymes which concern closely the improvement of fruit quality. The base sequence coding cDNA of sorbitol dehydrogenase which converts sorbitol of translocating sugar in Rosaceae fruit to fructose was determined in apple fruit. In tomato fruit, cDNAs of 6 isozymes of ACCsynthase were isolated and the expressions were investigated using the specific antibody for each isozyme. From melon fruit, the cDNA encoding ACCsynthase and ACCoxidase were isolated, the base sequences were determined and the gene expression were investigated with fruit development. The base sequence of cDNA encoding sucrose phosphate synthase, sucrose synthase and acid invertase concverning the biosynthesis and degradation of sucrose of translocating sugar in Citrus were determined. The gene encoding cysteine protease which functions the maturation and senscence was isolated from French bean seed and Vigna mungo seedling, and the structure of promotor and the gene expression were investigated. alpha-Arabinofurano sidase concerning the softening of Japanese pear fruit was purified.The second group tried to establish the plant-regeneration system and gene transfer method to for the transgenic fruit tree. In apples, the plant-regeneration from the callus of apple cultivar and the gene transfer via Agrobacterium became possible, and we succeeded the transfer of S6PDH chimeric gene. In persimmon, the plant was regenerated from callus and protoplast, and the recombinant of marker gene was formed through Agrobacterium infection. Further, in grape and citurs belonging sub-family of Rutaceae, we succeeded to get the embryogenic callus and to form the recombinant of GUS gene by co-culturing the callus with Agrobacterium. As described above, we could collect some genes concerning closely the fruit quality, and establish the plant-regeneration method and the gene trans fer method in main fruits crops in Japan.

该项目由两个小组进行。第一组尝试了与果实品质改良密切相关的几种酶的纯化和克隆。测定了苹果果实中山梨醇脱氢酶cDNA的碱基序列。从番茄果实中分离出ACC合酶6种同工酶的cDNA，并利用每种同工酶的特异性抗体研究了它们的表达。从甜瓜果实中分离ACC合成酶和ACC氧化酶的cDNA，测定其碱基序列，并研究其在果实发育过程中的表达。测定了柑橘蔗糖磷酸合成酶、蔗糖合成酶和酸性转化酶的cDNA序列。从菜豆种子和豇豆幼苗中分离到了半胱氨酸蛋白酶基因，并对其启动子结构和基因表达进行了研究。第二组研究了转基因梨树的植株再生体系和转基因方法。在苹果中，从苹果品种的愈伤组织再生植株和农杆菌介导的基因转移成为可能，并成功地将S6PDH嵌合基因转入苹果。以柿为材料，通过愈伤组织和原生质体再生植株，并通过农杆菌侵染形成标记基因重组体。在此基础上，以芸香科葡萄和柑橘为材料，通过与农杆菌共培养的方法，成功地获得了胚性愈伤组织，并形成了GUS基因的重组体。本研究为在日本主要水果作物中收集与果实品质密切相关的基因，建立植株再生方法和转基因方法奠定了基础。