|
Abstract Objective To construct eukaryotic expression vector of pcDNA3.1 (+)- FXYD5 gene and explore its effect on rat vascular smooth muscle cells(CRL- 1444). Methods The pBluescript II KS (+/- ) vector which contained ful- length FXYD5 cDNA was amplified by PCR, and cloned into pcDNA3.1 (+) plasmid. The recombinant plasmid was verified by PCR, double enzyme digestion analysis and DNA sequencing. CRL- 1444 cells were transfected with pcDNA3.1 (+)- FXYD5 using Lipofectamin 2000 as experimental group(PF), transfected with PCDNA3.1(+) as negative control group(NC) and not transfected as blank control group (BC). The expression of FXYD5 was measured quantitatively by real- time PCR. The effects of the gene on membrane Na+- K+- ATPase activity, proliferation and migration of CRL- 1444 cells were investi-gated. Results The FXYD5 gene was successful y cloned into pcDNA3.1(+). In comparison with BC group, the expres-sion of FXYD5 in PF group was increased by 17.312 (P<0.05). The membrane Na+- K+- ATPase activity was significantly higher in PF group than in BC group and NC group (P<0.05). The ability of cellproliferation and migration was significantly higher in PF group than in NC group (P<0.05). Conclusion The eukaryotic expression vector pcDNA3.1 (+)- FXYD5 is successful y constructed and FXYD5 expression increases in CRL- 1444 cells, which enhance the cellmembrane Na+- K+- ATPase activity, cellproliferation and migration ability.
|
|
|
|
|
|
