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Immunoassay of Phytochrome A Content in Photoperiod-sensitive Genic Male-sterile Rice

免疫测定光敏核不育水稻中光敏色素Ⅰ含量
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摘要 A rice mutant (Oryza sativa L. Nongken 58S) 'photoperiodsensitive male-sterile rice' (PGMR) is sterile under long day-length (LD), but fertile in short days-length (SD). Phytochrome (Phy) has proved to be the photoreceptor involved in photoperiod-induced male sterile process in PGMR.To speculate how phytochrome functions, PhyA levels in leaf extracts of PGMRand wild-type rice (Oryza sativa L. Nongken 58) under different photoperiods were measured by means of a sandwich enzyme-linked immunosorbent assay (ELISA). The top two leaves of treated rice plants were sampled at the end of the last dark phase for 10 cycles at photoperiod-sensitive stage for fertility alteration in the mutant. Compared with LD treatment, the SD treatment resulted in 39% increase of relative PhyA content (per unit fresh weight) in PGMR, but only in 18% increase in its wild-type. In addition, after 10 cycles of end of far-red irradiations of day,the heading and flowering dates of PGMR were delayed for 2 days, while its male fertility almost remained unchanged. It is well known that the end of day far-red responses are mediated by PhyB.These results suggested that in PGMR PhyA took part in regulation of male sterility, hut PhyB was involved in mediating flowering time. 光敏核不育水稻(农垦58S)是中国水稻研究中的一个重要发现,在水稻杂交育种上有巨大潜力。研究表明:农垦58S的雄性不育性受光周期调控,光敏色素是育性转变过程中光周期反应的光受体,然而,鉴于高等植物中存在至少两种不同的光敏色素分子,并且它们同时存在于水稻中,人们对调节农垦58S雄性不育过程的光敏色素分子种类及作用方式仍然不清楚。本文采用酶联免疫吸附测定法(ELISA),比较了不同光周期下农垦58S和对照品种农垦58叶片(光周期感受器官)中光敏色素I(phyA)的含量。从农垦58S的二次枝梗原基分化期开始,进行10天的光周期处理。一组为短日照(SD),另一组为长日照(LD)。在第10个暗期结束前,于暗绿光下收获每株水稻的最上部两片新展叶,立即保存在液氮中。样品在研钵中磨成液氮粉,然后加入提取液(含50mmol/LTris-HCI和0.2mol/L巯基乙醇,pH8.5)匀浆。粗提液经0.5%聚乙烯亚胺(PEI)沉淀后,上清液供ELISA分析。以燕麦phyA的多克隆抗体、单克隆抗体分别作ELISA的一抗和检测抗体。采用ELISA可以专一性地检测水稻phyA(Table1)。几次独立的实验说明,光周期处理对phyA含?
出处 《Developmental and Reproductive Biology》 1996年第2期51-59,共9页 发育与生殖生物学报(英文版)
关键词 PHYTOCHROME ELISA Male sterility Oryza sativa L. 水稻 光敏核不育 光敏色素I 测定 ELISA
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