FLAG-PCNA野生型和K164A突变型重组质粒的构建和表达
onstruction and expression of FLAG-PCNA wild type and K164A mutation plasmids
目的：构建pCMV-FLAG-PCNA野生型（Wt）和K164A突变型重组质粒，进行真核细胞表达和蛋白差异比较。方法：利用PCR扩增和定点突变技术，从质粒模板中分别扩增野生型和含K164A突变位点的目的基因全长片段，将其分别插入pCMV-N-FLAG真核表达载体中，构建PCNA重组质粒；然后，利用转染试剂将重组质粒导入293T细胞进行表达，比较野生型和突变型目的蛋白的表达差异。结果和结论：成功构建并正确表达了pCMV-FLAG-PCNA野生型（Wt）和K164A突变型重组质粒，但二者在真核细胞中的表达无明显差异，这为进一步研究PCNA相关结构功能奠定了基础。
Objective：To construct and express the pCMV-FLAG-PCNA wild type and K164A mutation plasmids. Methods：Target genes amplified through PCR and Site-directed Mutagenesis were inserted into the eukaryotic expression vector pCMV-N-FLAG. Then, the recombinant plasmids were transfected into 293T cell line to compare of their expression. Results and Conclusion: FLAG-PCNA wild type and K164A mutation plasmids were constructed and expressed succeedly. However, the difference of their expression was not obvious between these two recombinant plasmids. These results lay the foundation for further study of the PCNA structure and function.
淡松松、秦焕焕、刘朗夏、高学娟、刘小会
分子生物学细胞生物学遗传学
PCNAPCNA K164A重组质粒表达
PCNAPCNA K164ARecombinant plasmidsExpression
淡松松,秦焕焕,刘朗夏,高学娟,刘小会.FLAG-PCNA野生型和K164A突变型重组质粒的构建和表达[EB/OL].(2012-02-29)[2025-04-27].http://www.paper.edu.cn/releasepaper/content/201202-1124.点此复制
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