Sequence Analysis Of S Gene And Construction Of Full-length CDNA Clone Of Porcine Deltacoronavirus

Porcine deltacoronavirus(Porcine deltacoronavirus,PDCoV)is a new type of porcine intestinal coronavirus that has emerged in recent years.It was first reported in Hong Kong,China in 2012.In2014,the PDCoV epidemic broke out in Ohio,the United States.Subsequently,the disease was reported in China,Canada,Japan,South Korea,Thailand,Mexico and other countries and regions.PDCoV can infect pigs of different ages.Newborn piglets are more susceptible.Its clinical symptoms are mainly diarrhea,vomiting,dehydration and death in severe cases,with a mortality rate of 30%-40% for newborn piglets.However,the pathogenic mechanism of PDCoV is not yet fully understood,and the lack of effective vaccines for prevention and control has brought huge economic losses to the pig industry.Therefore,the establishment of the PDCoV reverse genetic operation platform as soon as possible is necessary for the subsequent in-depth study of the structure and function of the virus genome,the pathogenic mechanism,and the development of new vaccines.In this study,we used the PDCoV epidemic strain CH/XJYN/2016 isolated and preserved in the laboratory as the parent strain,sequenced and analyzed the cytotoxic S gene sequences of different generations,identified the mutation sites of different generations of cytotoxic S gene.And provided specific operation sites for the subsequent use of reverse genetic technology to study the molecular mechanism of pancreatin dependence and reduced pathogenicity of PDCoV.Furthermore,through the construction strategy of vitro ligation,the full-length genome was divided into five fragments,and the five target fragments were ligated in vitro by T4 ligase.The full-length cDNA clone and full-length in vitro transcripts of the genome were successfully obtained.In conclusion,in this study,the full-length genome sequence of PDCoV CH/XJYN/2016 strain P10 generation and the sequence information of different generations of cytotoxic S gene were successfully obtained,and the full-length cDNA and vitro transcripts of PDCoV genome were further obtained by vitro ligation technology,which laid an important foundation for the establishment of subsequent reverse genetics operation platform and the study of pathogenic mechanism of PDCoV.