山羊地方性鼻内肿瘤病毒ENTV-2FJ株的分离纯化和全基因组序列分析

doi:10.3969/j.issn.1674-7968.2020.02.013

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Abstract Enzootic nasal adenocarcinoma (ENA) caused by Enzootic nasal tumour virus of goats (ENTV-2) has been reported in many provinces in China in recent years. In order to clarify the virology and genome-wide characteristics of ENTV-2FJ strain, In this study, the nasal fluid of the diseased sheep was collected, and the virus of ENTV-2FJ strain was purified by differential centrifugation combined with sucrose density gradient centrifugation, and then subjected to electron microscopic observation and SDS-PAGE; Seven cDNA fragments of ENTV-2FJ isolate were amplified by reverse transcription PCR (RT-PCR), the amplified products were cloned into pMD19-T plasmid vector and sequenced, the whole genome sequence of the ENTV-2FJ strain was obtained by splicing with DNAstar 7.1 software and the sequence was analyzed. After purification of the virus of ENTV-2FJ strain, pure virions can be observed under electron microscope. The virions were spherical, 90~110 nm in diameter, with envelope and spikes; By SDS-PAGE analysis, the structural proteins of the ENTV-2FJ strain showed 9 bands with molecular weights of 118, 79, 60, 51, 28, 20, 16, 13 and 11 ku, the genome of the ENTV-2FJ strain was 7 469 bp in length, exhibiting 95.1%, 95.4%, 95.1%, 94.9% and 95.4% homology with ENTV-2SC, ENTV-2Shaanxi, ENTV-2Shaanxi2, ENTV-2Shaanxi3, ENTV-2Shaanxi4, respectively. Phylogenetic analysis based on the nucleotide sequence of ENTV-2 whole genomes, gag and env genes revealed that the ENTV-2FJ strain was in the same evolutionary branch as ENTV-2SC, ENTV-2Shaanxi, ENTV-2Shaanxi2, ENTV-2Shaanxi3, and ENTV-2Shaanxi4. These results lay the foundation for the establishment of the ENTV-2 assay and genetic variation analysis.

Key words： Enzootic nasal tumor virus of goats (ENTV-2) Purification Electronic microscope observation SDS-PAGE Genomes sequencing

Received: 24 July 2019

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Corresponding Authors: *hql562713@163.com

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	JIANG Jin-Xiu
	LIN Yu-Sheng
	ZHANG Jin-Peng
	YOU Wei
	HUANG Li-Li
	MAO Kun-Ming
	HU Qi-Lin

Cite this article:

JIANG Jin-Xiu,LIN Yu-Sheng,ZHANG Jin-Peng, et al. Isolation, Purification and Genome Sequence Analysis of Enzootic nasal tumor virus of Goats (Capra hircus) Strain ENTV-2FJ[J]. 农业生物技术学报, 2020, 28(2): 313-324.

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http://journal05.magtech.org.cn/Jwk_ny/EN/10.3969/j.issn.1674-7968.2020.02.013 OR http://journal05.magtech.org.cn/Jwk_ny/EN/Y2020/V28/I2/313

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