人核糖体40S亚基RPS11蛋白的原核表达和纯化

RPS11是核糖体小亚基40S的组成部分,属于核糖体蛋白S17p家族,由RPS11基因所编码,主要存在于真核生物中。本研究通过PCR扩增RPS11基因,构建pET43a-RPS11和pGEX-4T-1-RPS11原核表达载体。将重组质粒转入E.coli DH5α,序列测定正确后,将其转入E.coli BL21(DE3),加入IPTG诱导表达。表达蛋白经SDS-PAGE分析后,利用亲和层析法纯化蛋白。结果成功克隆RPS11基因,构建了原核表达载体pET43a-RPS11和pGEX-4T-1-RPS11,转化宿主菌E.coli BL21(DE3)中进行了表达。SDS-PAGE分析证实表达目的蛋白正确。通过Ni-TNA和GSH-Sepharose层析柱获得纯化蛋白,为进一步研究该蛋白的生物学功能奠定了基础。

Prokaryotic Expression and Purification of the Protein of Human Ribosomal 40S Subunit RPS11

RPS11 is a part of the small ribosomal subunit 40S,belonging to the family of ribosomal protein S17p,encoded by RPS11 gene,and mainly exists in eukaryotes.RPS11 gene was amplified by PCR,inserted pET43.1 a （＋） and pGEX-4T-1 to construct pET43a-RPS11 and pGEX-4T-1-RPS11 prokaryotic expression vectors.The recombinant plasmids were transformed into E.coli DH 5α,the vectors with correct sequence were transferred into E.coli BL21 （DE3）,and induced with IPTG.The expression of the protein was analyzed by SDS-PAGE,and the protein was purified by affinity chromatography.RPS11 gene was successfully cloned,prokaryotic expression vector,pET43a-RPS11 and pGEX-4T-1-RPS11 were conducted and expressed in transformed host strain E.coli BL21 （DE3）.SDS-PAGE analysis confirmed the target protein was expressed correctly.The protein was successfully purified by Ni-TNA and GSH-Sepharose column,which is very important for further study of the biological function of the protein.