改良菌落PCR快速鉴定TRIM28基因阳性重组子的方法研究  

An improved bacteria PCR technique for rapid screening TRIM28 gene positive recombinant

作　　者：梅志强[1] 张连美[1] 于海清[1] 段承刚[1] 杨曼曼[1] 傅俊江[1] 

机构地区：[1]泸州医学院医学基础研究中心,四川泸州646000

出　　处：《泸州医学院学报》2012年第3期241-244,共4页Journal of Luzhou Medical College

基　　金：国家自然科学基金项目(30371493;81172049)

摘　　要：目的:建立简便、可靠基因克隆菌落阳性重组子的筛选方法。方法:应用普通菌落PCR和改良的菌落PCR方法对TRIM28基因的阳性重组体进行阳性重组子的分析比较,同时探讨了PCR反应体系中模板浓度和DMSO对于扩增效率的影响。并对鉴定结果为阳性的克隆进行了进一步酶切和蛋白诱导表达分析。结果:运用改良的菌落PCR法简单快捷,结果可靠,反应体系中DMSO的浓度为4%左右可以很好地促进GC富集DNA模板的PCR扩增。其方法的可靠性得到了酶切验证和重组蛋白表达证实。结论:应用改良方法可以用于快速有效地筛选阳性重组菌落。Objective： To develope a simple and reliable method for recombinant colonies screening. Methods： Regular colony PCR and improved PCR method of the positive recombinant TRIM28 gene were analyzed and compared with, and the concentrations of the DNA template and DMSO for the efficiency of PCR reaction were analyzed. The results of identification for the positive clones were further tested by enzymatic digestion and the protein-induced expression. Results： The modified colony PCR method described was simple, fast and reliable. DMSO concentration of 4% could get the best result in the PCR reaction system for GC rich DNA template. The reliability of the method was verified by the enzymatic digestion and the protein-induced expression. Conclusion： The modified method can be used for screening the positive recombinant colonies fast and effectively.

关 键 词：菌落 PCR 改良 TRIM28 

分 类 号：Q785[生物学—分子生物学]