沙梨PpGAI3基因的分离与原核表达分析

发布时间：2018-01-26 01:09

  本文关键词： 沙梨 休眠 DELLA蛋白 PpGAI 原核表达　出处：《果树学报》2017年02期 　论文类型：期刊论文

【摘要】：【目的】分离沙梨DELLA蛋白编码基因PpGAI3,探究其蛋白在花芽休眠中的表达情况。【方法】从不同需冷量沙梨品种‘蜜雪梨’和‘黄花梨’中分离PpGAI3基因,重组到p EASY-Blunt E1表达载体中,转化BL21(DE3)宿主菌后经IPTG诱导表达,并对表达菌株进行SDS-PAGE和Western blotting检测。【结果】获得2种沙梨的PpGAI3基因,分别命名为PpGAI3mx和PpGAI3hh,Gen Bank登录号为KU954535和KX078215,其开放阅读框(ORF)为1 641 bp,编码546个氨基酸,预测其为亲水性不稳定蛋白。进化树分析表明PpGAI3mx和PpGAI3hh与白梨的亲缘关系最近;成功构建了p EASY-Blunt E1-PpGAI3原核表达载体,得到蛋白分子质量约60 ku的诱导表达产物。【结论】所得沙梨DELLA蛋白编码基因PpGAI3为植物DELLA蛋白家族中的成员,在梨花芽休眠过程中具有特异表达,且表达产物大部分以不溶性包涵体形式存在。
[Abstract]:[objective] to isolate the DELLA protein encoding gene PpGAI3. To investigate the expression of its protein in flower bud dormancy. [methods] PpGAI3 gene was isolated from the varieties' Honeyue 'and' Huanghua pear'. The expression was induced by IPTG after recombinant into the expression vector of p EASY-Blunt E1 and transformed into BL21DDE3. SDS-PAGE and Western blotting were used to detect the expression strain. [results] the PpGAI3 gene of two kinds of pear was obtained. The login numbers for PpGAI3mx and PpGAI3hhlgen Bank are KU954535 and KX078215, respectively. Its open reading frame (ORF) was 1 641 BP, encoding 546 amino acids. Phylogenetic tree analysis showed that PpGAI3mx and PpGAI3hh had the closest phylogenetic relationship with white pear. The prokaryotic expression vector of p EASY-Blunt E1-PpGAI3 was successfully constructed. The protein molecular weight was about 60 ku. [conclusion] the DELLA protein encoding gene PpGAI3 is a member of the plant DELLA protein family. There was a specific expression in pear bud dormancy, and most of the expressed products existed in the form of insoluble inclusion body.
【作者单位】： 福建农林大学园艺学院·福建农林大学园艺植物天然产物研究所;
【基金】：国家自然科学基金(31501694) 高等学校博士学科点专项科研基金(20113515110011)
【分类号】：S661.2
【正文快照】： 沙梨(Pyrus pyrifolia)为福建省主要落叶果树,具有冬季芽休眠的有益特性,喜温暖湿润环境,适应能力强,休眠期短,在我国南方地区被大面积推广[1]。近年来在引进‘翠冠’‘翠玉’等新品种进行更新换代的同时,仍缺少短低温梨优良品种,制约了福建亚热带气候对梨生产优势的发挥[2-3]，

本文编号：1464221