摘要
目的:构建Tudor-SN蛋白的Serine426(S426)、Serine781(S781)、Threonine240(T240)和Threonine429(T429)的点突变质粒,并使该重组质粒能够在He La细胞中融合表达。方法:利用定点突变技术,对Tudor-SN蛋白进行S426A、S781A、T240A、T429A点突变,通过双酶切的方法获得Tudor-SN.Mutants片段,最后将其连入到p CMV-N-Flag载体中。在He La细胞中转染该质粒,利用Western blot技术检测质粒表达效率。结果:(1)重组质粒经双酶切鉴定,可以观察到载体与Tudor-SN.Mutants的条带。(2)转染突变质粒后可看出He La细胞中有Flag蛋白表达。结论:质粒构建成功,可以用于下一步科学研究使用。
Objective： To construct eukaryotic Flag expressing recombinant pCMV-N-Tudor-SN.Mutants-Flag. Methods：The Serine426 （S426）, Serine781 （S781）, Threonine240（T240）and Threonineg29（T429） of Tudor-SN were transformed into Alanine by site- directed mutagenesis technique. Then the Tudor-SN.Mutants were obtained by restricting double enzyme digestion, and then inserted into pCMV- N-Flag vector. The recombinant plasmids were transfected into HeLa and observed by Western blot. Results： （ 1 ） The vector and Tudor-SN. Mutants could be observed by restricting double enzyme digestion. （2） Flag was expressed by HeLa which was transfected with recombinant plasmid. Conclusion：The recombinant plasmids of pCMV-N-Tudor-SN.Mutants-Flag are constructed successfully, and may be useful for further study.
出处
《天津医科大学学报》
2016年第1期5-8,共4页
Journal of Tianjin Medical University
基金
国家杰出青年基金资助项目(31125012)
教育部"创新团队发展计划"(IRT13085)
国家自然科学基金资助项目(31170830
31370749
31571380)
天津市应用基础与前沿技术研究计划(青年基金项目)(15JCQNJC09900)
