摘要
ESPRG3基因是一个与干细胞功能相关的基因,mESPRG3和hESPRG3基因被构建到原核及真核表达载体进行表达,原核表达蛋白用于制备多克隆抗体开展ESPRG3基因功能研究.真核表达载体EGFP被用于亚细胞定位.构建的mESPRG3基因原核及hESPRG3真核表达载体,成功进行了表达.利用IPTG诱导表达ESPRG3蛋白制备了多克隆抗体,免疫印迹和免疫组化结果表明制备的抗体具有特异性;表达的蛋白检测到体外具有结合DNA的能力.染色质免疫沉淀联合芯片技术检测到ESPRG3可以特异性结合到染色体上,且这些结合位点与Alu、及绝缘子序列具有关联性,可能通过这些位点调控胚胎干细胞的功能.
ESPRG3 is a gene related to hES cells.In order to investigate the function of ESPRG3,the mESPRG3 prokaryotic and hESPRG3 eukaryotic expression vector were constructed and polyclonal antibody was prepared.Molecular cloning and IPTG inducing were used to express and purify protein of ESPRG3 for the antibodies preparation.Western blot and imunohischemistry were used to identify the specificity of the antibody of Borealin.The recombinant proteins of ESPRG3 were found to show ability of binding DNA in vivo and in vitro with the ChIP-Chip assay and DNA select assay.These binding sites are overlapped with Alu and insu-lator elements.The function of it in ES cells may relate to these binding sites.
出处
《生命科学研究》
CAS
CSCD
北大核心
2012年第4期307-313,323,共8页
Life Science Research
基金
湖南省科技计划项目(2010FJ3026)
