Construction and identification of PC12 ceIIs with overexpressed CBS
YIN Weilan
LIAO Zhiqiang
YANG Bocheng
JIANG Luoyong
WEN Fang
WU Lixiang
Abstract：Objective To establish stable PC12 cell line with overexpression of cystathionine-β-synthase (CBS)and to identify it.Methods CBS primers were designed according to cDNA sequence of CBS gene from GenBank.The CBS gene was amplified using PCR and ligated into the lentiviral vector GV341.293T cells were transfected with the recombi-nant vector and two kinds of packaging viral vector.Viral supernatant was collected and was used to transfect PC12 cells. After puromycin screening,PC12 cells with CBS gene transfection were constructed.The CBS mRNA and protein of the PC12 cells and the empty vector transfected PC12 cells was detected by real-time fluorescent quantitative PCR and Western blotting,respectively.Results The CBS mRNA expression in the observation group,control group and empty vector group was 162.2 ±11.2,1.0 ±0.03 and 2.3 ±0.07,respectively.The CBS protein expression in the observation group,control group and empty vector group was 1.53 ±0.09,1.00 ±0.10 and 0.96 ±0.10,respectively.The CBS mRNA and protein expression levels of the observation group were higher than those of the control group and empty vector group (P <0.05). The difference of CBS mRNA and protein expression between the control group and empty vector group was not statistically significant.Conclusion The PC12 cell line with overexpressed CBS is successfully constructed.
Keywords：PC12 cellscystathionine-β-synthaselentiviral vector
Publication Date：2016-01-01
Online Publishing Date：2025-08-15(First online date of this platform, not the publication date of the document)
Pages：3( 17-19 )