Construction of shRNA expression plasmid and its inhibitive effect on HBV replication in vitro

ObjectiveTo construct small hair RNA(shRNA)expressiion vector specifically targeting the X and P gene sequence of hepatitis B virus(HBA)and transfecting the vector into infection cell model by nanometer polymers,and to evaluate its inhibitory effcet on the replication of HBV.MethodsThe interfering RNA sequences targeting HBV genetic coding region were designed and three pairs of oligonucleotide(64 nt)were synthesized.Then they were inserted into pRNAT-U6 to construct recombinant interfering plasmids S1,S2 and S3.The three recombinant plasmids were transfected into HBV infected HepG2 cells by nanometer transfection reagent.HBV mRNA was measured with real time PCR.ResultsThe replication of HBV was suppressed by the inter-fering plasmids S1 and S2 sigificantly.Compared to blank control,HBV mRNA of groups S1 and S2 were reduced to 70.2% and 61.2%,respectively(P<0.05).Unrelated control plasmid had no inhibitory effect.ConclusionThe shRNA expression vectors targeting HBV genetic coding region were successfully constructed and the study results show that they can suppress HBV mRNA in vitro.