摘要
为研究水稻开花调控的分子网络,构建了水稻品种中花11幼穗分化前倒二叶叶片的酵母双杂交cDNA文库。制备了高质量的总RNA,用Oligo(dT)引物反转录合成cDNA第一链和Long Distance PCR合成双链cD-NA,通过SMART同源重组交换技术在酵母菌株AH109中建立酵母双杂交所需的水稻倒二叶叶片cDNA文库。文库质量检测结果表明:cDNA文库的转化率为1.178×106/3μg pGADT7-Rec,文库滴度为2.65×108 cfu/mL,重组率为94.7%,插入片段长度为350～2 000bp。该文库可用来筛选水稻抽穗期基因的互作蛋白。
A yeast two-hybrid cDNA library of penultimate leaf harvested from the rice cultivar Zhonghua11 before its flowering transition was constructed to study the molecular network of flowering regulation. The high-quality total RNA was isolated and the first-strand cDNA was obtained by reverse transcription using primer Oligo（dT）. The double-strand cDNA was amplified by using Long Distance PCR. The cDNA library of rice penultimate leaf was generated by using homologous recombination-mediated SMART technology in yeast strain AH109. The testing results of library＇s quality showed that the transformation efficiency of cDNA library was 1. 185× 10^6 transformants/3 μg pGADTT-Rec with the titers of 2.65 × 108 cfu/mL and the recombinant rate of 94. 7%. The sizes of inserted fragments were ranged from 350 bp to 2 000 bp. These data indicated that the cDNA library could be used to screen in- teracting proteins of the flowering time gene in rice.
出处
《华中农业大学学报》
CAS
CSCD
北大核心
2012年第4期404-409,共6页
Journal of Huazhong Agricultural University
基金
国家自然科学基金项目(30970172)
作者简介
彭强，硕士研究生．研究方向：水稻功能基因组学．E—mail：450058876@163．com
通讯作者：吴昌银，博士，教授．研究方向：水稻功能基因组学．E-mail：cywu@mail．hzau．edu．cn
