RESISTANCE OF SIGCRV TRANSGENIC RARE MINNOW(GOBIOCYPRIS RARUS)TO GRASS CARP REOVIRUS

In this study,three shRNA expression vectors pH1siGCRV(x)-CMVeGFP were constructed by employing grass carp H1 promoter,using outer capsid protein VP7 gene of grass carp reovirus (GCRV) as target gene and eGFP as report gene.CIK cells transfected experiment suggested that pH1siGCRV2-CMVeGFP had the strongest suppression on GCRV replication.Transgenic P0 group of fish were obtained by microinjecting pH1siGCRV2-CMVeGFP to rare minnow(Gobiocypris rarus)embryos.The transgenic fish had only 30% mortality after GCRV infection.It was shown that the resistance of the transgenic fish to GCRV has significantly improved.Moreover,the results of real-time PCR on VP7 gene in spleen,hindgut and liver confirmed that the content of GCRV in transgenic fish was significantly lower than control and gradually reduced over time,indicating that the replication of GCRV has been suppressed in transgenic fish.This research has provided a solid foundation for producing GCRV-resistant transgenic fish.