Screening And Identification Of The Zinc Finger Nuclease And TALEN Targeting Porcine Sn Gene

Porcine reproductive and respiratory syndrome is one of the most important diseases threatening the global pig industry. Porcine reproductive and respiratory syndrome virus (PRRSV) is recognized with great virulence difference, high mutation rate and low antigenicity, thus it is difficult to obtain strong immune protection with the existing vaccines. PRRSV mainly infects macrophages, and which is mediated by sialic acid adhesin (Sialoadhesin, Sn) receptors and CD163receptors. Compared with viral proteins, virus receptors on target cells is highly conserved, so targeting virus vectors may block the infection of different virus strains, and these technologies have become the new anti-viral infection strategies. Zinc finger nuclease (ZFN) technology is the recently developed gene modification technology, while the transcription activator-like effector nuclease (TALEN) technology is the newest gene designated modified technology.In order to obtain ZFN and TALEN targeting porcine Sn for transgene study in vivo, this study designed5pairs of ZFN and1pair of TALEN targeting porcine Sn. In vitro transcription/translation system and E.coli expression system was employed respectively to express the nuclease to digest the plasmid DNA containing the target sequence. Then a specific cleavage activity ZFN or TALEN can be screened. The results show that these five pairs ZFN can not effectively cut the target sequence, but the TALEN can cut the supercoiled plasmid containing the target sequence. These results further show that the use of existing bioinformatics software to design and screen effective ZFN is blind and inefficient; While TALEN can be designed more effectively.