重庆医学Issue(10)：1164-1167,4.DOI:10.3969/j.issn.1671-8348.2014.10.004
SIGIRR胞内区酵母双杂交诱饵质粒的构建及自激活作用检测
Construction of bait plasmid containing SIGIRR cytoplasmic tail and detection of self-activation in yeast two hybrid system
摘要
Abstract
Objective To construct the bait plasmid of pSos-single immunoglobin IL-1 receptor related protein (SIGIRR) in Cy-toTrap yeast two hybrid system ,and to test its self-activation .Methods The cDNA fragments of SIGIRR(480 -1 230 bp) were amplified from pReceiver-LV19-SIGIRR and ligated into the bait plasmid pSos to generate the plasmid pSos-SIGIRR .The pSos-SI-GIRR was identified by DNA sequencing and dual-site endonuclease digestion .Then the recombinant plasmid and control plasmid were introduced into the yeast cell cdc25H .The transformants were inoculated on plates of 25 ℃ /SD/Glucose(-UL) ,25 ℃/SD/Ga-lactose(-UL) ,37 ℃ /SD/Glucose(-UL) and 37 ℃ /SD/Galactose ,respectively and the proliferation ability of transformant was ob-served for 6 d .The Western blot was adopted to detect the expression of target protein .Results The pSos-SIGIRR vector was cor-rectly constructed and proved of no self-activation and toxic action .The Western blot showed that the target protein was expressed in a form of fusion protein of 170KD .Conclusion The bait plasmid containing SIGIRR cytoplasmic tail can be applied to the yeast two-hybrid system and lays the important foundation for seeking the interacting protein with SIGRR from the human lung cDNA li-brary in .关键词
免疫球蛋白白细胞介素1受体相关蛋白/双杂交系统技术/诱饵质粒/自激活Key words
single immunoglobin interleukin-1 receptor related protein/two-hybrid system techniques/bait plasmid/self-activation引用本文复制引用
陈旭昕,冯华松,段蕴铀,吴学玲,钱桂生..SIGIRR胞内区酵母双杂交诱饵质粒的构建及自激活作用检测[J].重庆医学,2014,(10):1164-1167,4.基金项目
国家自然科学基金资助项目（30600272）。 （）
