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Construction and Screening of Plasmid Expression Vectors Encoding the Short Hairpin RNA Targeting DNA Polymerase of Goatpox Virus
An-Tao Wang1,3, Chuang-Fu Chen1*, Jun Qiao1*, Hui Zhang2, Na Zhang1, Sheng-Wei Hu2
1College of Life Science, Shihezi University, Shihezi 832003, China;2College of Animal Science & Technology, Shihezi University, Shihezi 832003, China; 3Laboratory of Xinjiang Endemic and Ethnic Disease, Shihezi 832003, Ch
Abstract: According to the gene sequence of DNA polymerase of Goatpox virus， 5 specific siRNA were synthesized， annealed and cloned into RNAi-Ready pSIREN-RetroQ ZsGreen vector. The recombinant plasmids were transformed into E.coli DH5α and identified by sequencing. The siRNAs were transfected into the BHK-21 cells. After infection experiment， siRNA vectors were extracted， and evaluated inhibiting effect of siRNA vectors on the gene of DNA polymerase of the Goatpox virus by real-time quantitative PCR (Q-RT PCR). The results showed that the genes of the DNA polymerase of the Goatpox virus could be inhibited by siRNA vectors with an inhibition rate of 63%~93.8%， as well as inhibition rate of pSI-W2 was the obvious highest and 93.8%. In this study， interference sequences， which were able to inhibit the multiplication of Goatpox virus in vitro， obviously， were screened from BHK-21 cells by RNAi technology， and can facilitate further study on the anti-Goatpox virus transgenic sheep.
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