Construction of the eukaryotic expressing vector of human PAK2 gene and the expression and location in gastric cancer cells

Objective To construct an eukaryotic expressing vector of human PAK2 gene and identify its recombinant plasmid expression and location in the gastric cancer cells.Methods Total mRNA was extracted from HeLa cells,cDNA was formed by reverse transcription.The PAK2 coding sequence was amplified by polymerase chain reaction(PCR)method and cloned into pcDNA3.1A vector.After the target region was sequenced,the plasmid was transfected into SGC-7901 cell lines.The expression of the pcDNA3.1A-PAK2 recombinant plasmid in SGC-7901 cells was proved by Western blot.The location of the plasmid in the cells was observed by using laser scanning confocal microscopy.Results hPAK2 was successfully constructed into pcDNA3.1A expressing vector.The length of the fragment was about 2000 bp,identified by restruction enzyme digestion.The expression of pcDNA3.1A-PAK2 recombinant plasmid in SGC-7901 cells was detecteded by Western blot,with a molecular weight of about 75 kDa.The plasmids were localized more in the cytoplasm,less in the nucleus.Conclusion The hPAK2 full length sequence was successfully cloned into eukaryotic expressing vector,the recombinant plasmids were localized mainly in the cytoplasm.