Cloning,Phylogenetic And Structural Analysis Of P32,GPCR And RPO30 Genes Of The First Chinese LSDV Isolate

Lumpy skin disease(LSD)is an acute or subacute viral infectious skin disease in bovine caused by lumpy skin disease virus(LSDV).The main clinical symptoms were fever(over 40?),skin nodules,swelling of superficial lymph nodes;it also has some other serious consequences such as the decrease of milk production,abortion,stillborn foetus,temporary or permanent sterility and death.Therefore,it is an exotic infectious disease that has a severe influence on the development of animal husbandry in China.In recent years,with the continuous outbreak and epidemic of LSD in other parts of the world,this disease was first reported in Yili Kazak Autonomous Prefecture of Xinjiang in August 2019 in China.Due to the lack of relevant research on LSDV etiology and molecular biology at present in China,this study cloned and sequenced the complete P32,GPCR and RPO30 genes of the strain,LSDV/Xinjiang/2019,in the State Key Laboratory of Veterinary Etiological Biology,Lanzhou Veterinary Research Institute Chinese Academy of Agricultural Sciences.The phylogenetic relationship and structural characteristics with other foreign isolates were also analyzed in this study.Objective: 1.To provide a reference for tracing LSDV by conducting the genetic evolution analysis of P32,GPCR and RPO30 genes of the LSDV/Xinjiang/2019 strain.2.To provide the reference for the development of LSD molecular diagnostic reagents and vaccines of LSDV through analyzing biological information of the proteins encoded by P32,GPCR and RPO30 genes respectively of LSDV/Xinjiang/2019 strain.Methods: 1.According to the CaPV gene sequences registered by GenBank,three pairs of specific primers were designed and synthesized to clone and sequence P32,GPCR and RPO30 genes of the LSDV/Xinjiang/2019 isolated strain.Then,t homology of nucleotide and genetic evolutionary relationship analysis wereconducted on the sequencing results with other CaPV isolates.2.This research predicts and analyzes the basic physicochemical property and structure of the P32,GPCR and RPO30 proteins by utilizing some bioinformatics methods and software,such as ExPASy ProtScale,SingleIP 5.0 and SOPAM.Results: 1.The specific fragments of P32,GPCR and RPO30 genes of the LSDV/Xinjiang/2019 strain were successfully amplified from cell cultures by using PCR,which is consistent with the expected results.The sequencing result has been uploaded to the GenBank(the registration numbers are MN598005,MN598006 and MN598007 respectively).The homology analysis has proved that the nucleotide consistency of the P32,GPCR and RPO30 genes of LSDV/Xinjiang/2019 strain and other isolated strains of LSDV is 88.3% ~ 99.8%,95.8% ~ 99.2% and 93.7% ~ 99.7%,respectively.The results of the phylogenetic trees suggest that LSDV/Xinjiang/2019 strain is more closely related to the isolated strain from Russia.2.The results of bioinformatics analysis show that the proteins encoded by P32,GPCR and RPO30 genes are composed of 322,381 and 202 amino acids,respectively;the main structures of the three proteins are ?-helix and random coil.Only GPCR protein is a hydrophobic protein,while the hydrophilic regions of the P32 protein and RPO30 protein are more abundant.The tertiary structure models of these three proteins with high matching degree with the template protein were predicted and screened out.Conclusions: 1.The complete genes of P32,GPCR and RPO30 of LSDV/Xinjiang/2019 strain,the first LSDV isolate in China has been successfully cloned,and the genetic evolution analysis was conducted,which provides a reference for the traceability of LSDV.2.The structure and function of P32,GPCR and RPO30 proteins of this strain have been predicted and analyzed for the first time,providing valuable information for the development of the new molecular vaccine and diagnostic reagents.