小反刍兽疫RT-PCR的鉴定及N基因序列分析.pdf

Asian Case Reports in Veterinary Medicine 亚洲兽医病例研究, 2015, 4(4), 31-38 Published Online October 2015 in Hans. /journal/acrpvm /10.12677/acrpvm.2015.44006 Identification of RT-PCR and Sequence Analysis of N Gene in Pest Des Petits Ruminants Virus 1,2 1 1 1 1 1 Xiaopeng Li , Hualiang Chen , Junqiong Zhai , Xia Zhou , Song Cheng , Wenjuan Zhang , Manlin Luo1*, Shaolun Zhai2* 1 College of Veterinary Medicine, South China Agricultural University, Guangzhou Guangdong 2 Animal Disease Diagnostic Center, Institute of Animal Health, Guangdong Academy of Agricultural Sciences, Guangzhou Guangdong * * Email: 710510116@, 280029676@ rd th th Received: Sep. 23 , 2015; accepted: Oct. 6 , 2015; published: Oct. 19 , 2015 Copyright © 2015 by authors and Hans Publishers Inc. This work is licensed under the Creative Commons Attribution International License (CC BY). /licenses/by/4.0/ Abstract Two pairs of primers were designed to amplify the partial P gene and full-length N gene based on their own conserved sequences, respectively. RNA of suspected samples with PPRV was extracted and amplified by RT-PCR, from which target fragments of 766 bp and 1578 bp were obtained. Meanwhile, the RT-PCR reaction conditions, including annealing temperature and sensitivity, were optimized and 850 copies of PPRV can be tested under these conditions. PCR product of N gene was cloned into pMD-19T vector and the recombinant plasmid was transformed into DH5αcompetent cells and sequenced, which showed that N gene wa