摘要：

AIM: To construct a eukaryotic expression vector containing short hairpin RNA(shRNA) targeting the maspin gene. METHODS: A shRNA targeting the maspin gene was ligated to the pGenesil-1.1 plasmid expressing green fluorescent protein. Recombinant maspin/pGenesil positive plasmid and YX/ pGenesil negative plasmid were constructed. After recombinant plasmids were transfected into MKN-28 cells, RT-PCR and Western blot were performed to detect maspin mRNA and protein expression, respectively. RESULTS: The recombinant plasmids were validated by gel electrophoresis and DNA sequence analysis. RT-PCR and Western blot analyses indicated that after successful transfection, both maspin mRNA and protein expression in the maspin/pGenesil group was dramatically down-regulated, compared with that in the YX/ pGenesil group(0.127 ± 0.02 vs 0.510 ± 0.01, P 0.05; 0.24 ± 0.10 vs 0.65 ± 0.09, P 0.05). CONCLUSION: The maspin shRNA recombinant plasmid has been constructed successfully. Transfection with the maspin shRNA recombinant plasmid could inhibit the expression of maspin mRNA and protein effectively in human gastric cancer cell line MKN-28.

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