Xu Xiao-jie, Fan Zhong-yi, Wang Ling-xue, Zhang Hao, Ding Li-hua, DU Nan, Ye Qi-nong
Beijing Institute of Biotechnology, Beijing 100850, China.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi. 2011 Aug;27(8):843-5.
To construct the prokaryotic expression vector of human histone-1, obtain the purified GST-H1 protein, and detect its activity.
Human histone-1 coding region was amplified from human mammary cDNA library, and was inserted into prokaryotic expression vector pGEX-KG. The recombinant plasmid pGEX-KG-H1 was transformed into E.coli Rossate. The expressed product was purified by GST-Sepharose 4B beads and identified by SDS-PAGE and Western blot analysis.
The DNA fragment of about 650 bp was successfully amplified by PCR, cloned into pGEX-KG, and identified by sequencing. The recombinant protein of about M(r); 52 000 was successfully induced, purified and tested well by Kinase assay.
The recombinant protein of GST-H1 is obtained successfully, which lay the foundation for further research on cell cycle control.
构建人组蛋白-1的原核表达载体，获得纯化的GST-H1蛋白，并检测其活性。
从人乳腺cDNA文库中扩增出人组蛋白-1编码区，将其插入原核表达载体pGEX-KG。将重组质粒pGEX-KG-H1转化至大肠杆菌Rossate。表达产物用GST-Sepharose 4B磁珠纯化，并用SDS-PAGE和Western blot分析进行鉴定。
通过PCR成功扩增出约650 bp的DNA片段，克隆至pGEX-KG中，并经测序鉴定。成功诱导、纯化出约M(r)为52 000的重组蛋白，且经激酶测定效果良好。
成功获得GST-H1重组蛋白，为进一步研究细胞周期调控奠定了基础。
