Live Attenuated Pseudorabies Virus And Herpes Simplex Virus Type 1 Developed Using The CRISPR/Cas9 System

Pseudorabies virus(PRV)and herpes simplex virus type 1(HSV1)are double strand DNA virus,belongs to herpes simplex virus?Pseudorabies,also known as Aujeszky's disease,will cause pseudorabies in pigs and pigs will eventually be death due to illness.Since the end of the 1970s,China imported Bartha-K61 strain for the prevention of Pseudorabies from Hungary and this disease can be well controled from 1990s to2010.However since the beginning of 2011,with the emergence of mutants virus,pseudorabies virus outbreak in bigs which ever have vaccinated with Bartha-K61 strains,bringing huge economic losses to the pig breeding industry.At present,traditional vaccine can not effectively prevent the outbreak of the virus in pigs and the preventive vaccine can't meet the demand of the market.So,it's urgent to development a new vaccines for the prevention of the disease.HSV1 will cause herpetic stomatitis,herpetic keratoconjunctivitis and herpes simplex encephalitis;in addition,HSV1 also have close relationship with Alzheimer's disease(Alzheimer,disease,AD)and Parkinson's disease(Parkinson 's disease,PD).Now,the vaccine for preventing HSV1 has not been successfully developed.It's have great significance that developing a effective vaccine of HSV1 to prevent the spread of HSV1 in the crowd.The CRISPR/Cas9 system have been developed in recent years and been used in the field of gene editing widely.We successfully knocked out the gE and TK gene of the PRV,as well as the gC and TK genes of HSV1 by using the system.After the purification with plaque,we have obtained three PRV attenuated strain,namely gE gene deletion strain(PRV-gE-),TK gene deletion attenuated strain(PRV-TK-),gE and TK double gene deletion strain(PRV-gE-/TK-)and two HSV1 strain,respectively gC gene deletion strain(HSV1-gC-),TK gene deleted attenuated Strain(HSV1-TK-).After the test of cell and animal,PRV-gE-?PRV-TK-?HSV1-gC-?PRV-gE-/TK-virulence is attenuated;HSV1-gC-virulence is stronger than HSV1-TK-but they are weaker than the wild type virus.With PRV-TK-,PRV-gE-/TK-,HSV1-gC-and HSV1-TK' to immune the mice,mice can resist the lethal dose of wild type virus infection.In the end,we think that PRV-TK-,PRV-gE-/TK-and HSV1-TK-can be used for production of vaccine candidate strain,But PRV-gE-and HSVl-gC-is not suitable as a vaccine candidate strains because of strong virulence.