Construction of eukaryotic expression plasmid pcDNA3.1-KISS-1 and its metastasis inhibiting effect to EC1 cells.

Objective To clone human esophageal tissue KISS-1 gene and construct its eukaryotic expression vector pcDNA3.1-KISS-1.To detect its expression in EC1 cells and observe the influence of transfection KISS-1 gene on the invasion and migration ability of EC1 cells.Methods Total RNA was extracted from human esophageal tissue.KISS-1 cDNA was isolated by using RT-PCR,and cloned into the eukaryotic expression vector pcDNA3.1.The eukaryotic expression plasmid pcDNA3.1-KISS-1 was verified by sequencing and enzyme digestion analysis.Using liposome-mediated transfection technique,the eukaryotic expression vector pcDNA3.1-KISS-1 was transfected into EC1 cells.The expression of protein in EC1 cells was detected by Western blot.The effect of KISS-1 expression on the invasion and migration of EC1 cells was investigated by Boyden-chamber system.Results pcDNA3.1-KISS-1 recombinant plasmid was successfully constructed and it had a higher level expressed in EC1 cells.Matrigel invasion and migration assay showed that the invasion and migration of cells transfected with KISS-1 were less than those in cells transfected with pcDNA3.1 and control cells(P0.05).Conclusion The expression of KISS-1 could inhibit EC1 cells invasion and migration in vitro.