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[The interference effect of lentiviral mediated shRNA on the expression of Pin 1 in A549 cells]
Shuai Zhao1, Wenbin Dong, Chan Zhang
1Department of Newborn Medicine, Affiliated Hospital, Luzhou Medical College, Luzhou 646000, China.
Objective:
To construct a Pin1shRNA lentiviral vector and transfect stably A549 cells.
Methods:
The peptidyl-proplyl cis-trans isomerase 1(Pin1) gene sequences from GenBank were cloned into the pLenR-GPH-shRNA lentiviral vector. The pLenR-GPH-shRNA and lentivector pakaging plasmid mix were cotransfected into human embryonic kidney cells HEK293T to package lentiviral particles. The virus supernatant was harvested, and then the virus titer was determined by serial dilution assay. The A549 cells were transduced with the constructed lentiviral vectors. Real-time quantitative PCR (qRT-PCR) and Western blotting were used to evaluate the Pin1 expression in different groups.
Results:
Restriction enzyme analysis and sequencing demonstrated that the recombinant plasmid named pLenR-GPH-Pin1shRNA was successfully prepared and had a high titer. qRT-PCR and Western blotting revealed that the significant down-regulation of Pin1 expression in A549 cells.
Conclusion:
The lentiviral mediated shRNA can effectively down-regulate the expression of Pin 1 in A549 cells.
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