The Whole Genome Sequencing And Semi-nested RT-PCR Detection Method Of Transmissible Gastroenteritis Virus HQ2016 Strain Was Established

Transmissible gastroenteritis virus(TGEV)is the etiological agent of transmissible gastroenteritis(TGE),which can cause vomit diarrhea with high morbidity in pigs of all ages,as well as high mortality in suckling piglets with high morbidity in suckling piglets with in two weeks nesrly 100%.TGE usually outbreaks in cold season and spring cause significant economic loss in swine-raising farms.The complete genome sequence of the clinically isolated strain TGEV HQ2016 was used Illumina next-generation sequencing of the TGEV HQ2016 strain.The total of 28 585 nt nucleotides were determined for TGEV HQ2016 strain,encompassing the5'-ORF(1a/1b)-S-3a-3b-sM-M-N-ORF7-3'.Percentage of nucleotide identity of TGEV HQ2016genome compare with others showed that share high identity with AYU,HX,WH-1,PUR46-MAD,Purdue.Phylogenetic analysis based on the complete genome sequence of TGEV HQ2016,other referenced strains showed that TGEV HQ2016 Strain had more closely related to purdue group than to Miller group.TGEV HQ2016 strain is closely related to the Purdue strain,and it is far away from the Miller group strain.Recombinant analysis of TGEV HQ2016 showed that there were not recombination regions in the TGEV HQ2016 strain.In order to establish an semi-nest RT-PCR method for the detection of TGEV,three specific primers were designed according to the TGEV HQ2016 strain and other TGEV strain sequence of N gene published by GenBank,the reaction system was established and optimized.The results showed that the method could successfully amplify the bands of 483 bp and 338 bp and had good specificity to TGEV,there is no cross reaction with PEDV,PRov,PBoV and PDCoV,and the lowest sensitivity was 1.86×10~-11 pg/?L.This semi-nest RT-PCR method has high sensitivity and specificity.The semi-nest RT-PCR showed the positive and can accurately diagnose TGEV infection rate was 36%in 50 samples of pig diarrhea,which provides an effective method for clinical detection and epidemiological investigation of TGEV.Genome property analysis and establish an semi-nest RT-PCR method for the detection of TGEV strain will not only contribute to the basic reseach of TGEV,but also assist in developing relevent biological products,the diagnosis and controlling of TGEV field strain.