Abstract
Objective To construct a stable over- expression lentivirus- mediated vector and transfect it into human gastric cancer cell line SGC7901 for investigating its effects on proliferation and migration of GC7901 cells. Methods ADP ribosylation factor guanylate kinase 1(ASAP1) gene coding region was cloned into lentivirus vector.Lentivirus particles were infected into the human gastric carcinoma cell line SGC7901 to upregulate the expression of ASAP1 gene.The up- regulated efficiency of targeting ASAP1 gene at mRNA level was detected by real- time quantitative polymerase chain reaction(Real- time PCR), the effect on proliferation of mesenchmal stem cells was assayed by 3-(4,5- dimethylthiazol- 2- yl)- 2, 5- diphenyltetrazolium bromide assay, and the migration ability was detected by Transwell motility assay. Results The lentiviral vector targeting ASAP1 gene was constructed successfully,and a stable human gastric cancer cell line SGC7901 line that up- regulated ASAP1 was established.Real- time quantitative PCR and Western blotting results showed that the expression of ASAP1 gene was efficiently up- regulated by infecting LV- ASAP1- GFP(P<0.01).255.4% of control group on mRNA expression and 290.2% of control group on protein expression.The 3-(4, 5- dimethylthiazol- 2- yl)- 2,5- diphenyltetrazolium bromide assay and growth curve showed that the over- expression of the ASAP1 gene successfully increased the proliferative capability of SGC7901 cells,with optical density values reaching 130.1%,136.0% and 149.1% of control group at 1,2,3 d respectively.The Transwell assay also showed similar increasing results on the migration ability(P<0.01). Conclusion The lenvivirus- mediated over- expression of the ASAP1 gene increases the proliferation and migration abilities of human gastric cancer cell line GC7901. Key words: Gastric carcinoma; ADP ribosylation factor guanylate kinase 1
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