pEGFP-C2-人类p100蛋白SN和TD片段质粒构建

Constructions of pEGFP-C2-p100-SN and pEGFP-C2-p100-TD recombinant plasmids

目的:分别构建含有人类p100蛋白SN片段和TD片段基因的真核绿色荧光蛋白表达载体pEGFP-C2-p100-SN和pEGFP-C2-p100-TD。方法:利用EcoRI和BamHI限制性内切酶对已构建好的质粒PSG5-p100-SN和PSG5-p100-TD进行双酶切以获得目的片段p100-SN和p100-TD基因片段,回收纯化后连接到质粒pEGFP-C2上。结果:通过对重组质粒进行菌落PCR鉴定以及酶切鉴定均能从PCR产物和酶切产物中观察到p100-SN和p100-TD片段。结论:成功构建了可以在真核细胞内表达绿色荧光蛋白和目的蛋白的融合蛋白的重组质粒pEGFP-C2-p100-SN和pEGFP-C2-p100-TD,可为有关人类p100蛋白功能及作用机制研究奠定基础。

Objective： To construct eukaryotic green fluorescence protein expressing recombinant plasmids, pEGFP-C2-p100-SN and pEGFP -C2-p100-TD, which contained human p100 Staphylococcal Nuclease （SN） domain and Tudor （TD） domain respectively. Methods： The p100-SN and p100-TD fragments were purified after PSG5-p100-SN and PSG5-p100-TD were digested by Eco RI and Barn HI. And then they were inserted into pEGFP-C2 fluorescent expressing vector respectively. Results： The fragments of p100-SN and p100-TD were detected in the products of the restriction enzyme digestion and PCR. Conclusion： The fluorescent expressing recombinant plasmids, which expressed p100-SN-GFP and p100- TD-GFP fusion proteins, are constructed successfully. The studies about the function of human p100 pro- tein are put forward.