基于RNA-Seq筛选ALV-J感染汶上芦花鸡肝差异表达致病相关基因

doi:10.11843/j.issn.0366-6964.2020.08.025

畜牧兽医学报 ›› 2020, Vol. 51 ›› Issue (8): 2003-2011.doi: 10.11843/j.issn.0366-6964.2020.08.025

基于RNA-Seq筛选ALV-J感染汶上芦花鸡肝差异表达致病相关基因

张会永¹, 吴井生², 杨剑波², 俞燕¹, 薛倩¹, 殷建玫¹, 朱云芬¹, 朱静¹, 苏一军¹, 李国辉^1*, 韩威^1*

1. 中国农业科学院家禽研究所, 江苏省家禽遗传育种重点实验室, 扬州 225125;
2. 江苏农林职业技术学院, 镇江 212400

收稿日期:2020-01-16 出版日期:2020-08-25 发布日期:2020-08-19
通讯作者: 韩威,主要从事家禽遗传资源保护、评价与利用研究,E-mail:hanwei830@163.com;李国辉,主要从事家禽遗传资源保护、评价与利用研究,E-mail:sahui2008@163.com
作者简介:张会永(1983-),男,河北承德人,本科,主要从事家禽遗传资源保护、评价与利用研究,E-mail:zhyong1983@163.com;吴井生(1979-),男,江苏泰兴人,副教授,博士,主要从事遗传育种研究,E-mail:17190607@qq.com
基金资助:
江苏农林职业技术学院科技项目（2017kj03）；江苏省六大人才高峰项目（NY-024）；江苏省属公益类科研院所自主科研经费项目（BM2018026-1）；江苏省重点研发计划（现代农业）专项（BE2019353）；江苏省家禽遗传育种重点实验室资助项目（JQSAB-ZZ-202007）；江苏省高校青蓝工项目

Application of RNA-Seq Technology for Screening the Differentially Expressed Pathogenic Genes in Wenshang Barred Chicken Liver Infected by Subgroup J Avian Leukosis Virus

ZHANG Huiyong¹, WU Jingsheng², YANG Jianbo², YU Yan¹, XUE Qian¹, YIN Jianmei¹, ZHU Yunfen¹, ZHU Jing¹, SU Yijun¹, LI Guohui^1*, HAN Wei^1*

1. Key Laboratory for Poultry Genetics and Breeding of Jiangsu Province, Poultry Institute, Chinese Academy of Agricultural Sciences, Yangzhou 225125, China;
2. Jiangsu Vocational College of Agriculture and Forestry, Zhenjiang 212400, China

Received:2020-01-16 Online:2020-08-25 Published:2020-08-19

摘要/Abstract

摘要： 为挖掘感染ALV-J汶上芦花鸡的肝差异表达致病相关基因。本研究以汶上芦花鸡为试验素材，分别在42和300日龄进行ALV血液病毒分离检测筛选ALV阴性和阳性个体，对ALV阳性个体有病理变化的肝组织和阴性个体肝组织进行PCR检测，分别选取3只ALV阳性（G1组）和阴性（G2组）个体的肝组织，并对G1组PCR产物测序、聚类分析确定ALV亚型，利用RNA-Seq测序技术筛选差异表达基因，并进行功能分析，利用荧光定量qRT-PCR对部分差异表达基因进行验证。结果表明，G1组样品经PCR检测、PCR扩增产物测序和聚类分析确定ALV为J亚型，转录组分析发现，共有42个差异基因在GO和KEGG中富集，其中，上调基因18个，下调基因24个。随机选取的5个差异表达基因qRT-PCR验证结果与RNA-Seq测序结果相一致。GO分析显示，差异表达基因主要涉及细胞过程、代谢过程、对刺激的反应、免疫系统等；KEGG分析显示，差异表达基因主要富集在细胞过程、信号传导、疾病、新陈代谢等信号通路。本研究通过转录组分析发现了影响ALV-J致病性的多个基因和关键信号通路，为深入了解ALV-J致病的分子机制提供了思路。

关键词: 汶上芦花鸡, J亚群白血病病毒, 转录组测序, 差异表达基因

Abstract: The aim of this study was to screen the differentially expressed pathogenic genes in liver of Wenshang Barred chicken infected by subgroup J avian leukosis virus(ALV-J). ALV uninfected and infected Wenshang Barred chickens were pre-screened by virus isolation from blood at 42 and 300 days of age. The liver tissue samples from both ALV infected chickens which had the pathological changes and ALV uninfected chickens were identified accurately by PCR detection,then the liver tissues from 3 ALV-positive (G1 group)individuals and 3 ALV-negative (G2 group) individuals were sampled for further study. PCR ampilification products of 3 ALV-positive individuals were confirmed as subgroup J by sequencing and cluster analysis. The liver tissue samples from both 3 ALV-J positive individuals and 3 ALV-negative individuals were selected for screening the differentially expressed genes by RNA-Seq technology,then functional analysis was conducted. The partial differentially expressed genes were verified by qRT-PCR. The results showed that 42 differentially expressed genes were enriched in GO and KEGG databases, which included 18 up-regulated genes and 24 down-regulated genes. Moreover, expressions of 5 differentially expressed genes randomly selected were verified by qRT-PCR, which confirmed that mRNA expression levels were consistent with RNA-Seq sequencing results. GO analysis results indicated that the differentially expressed genes were mainly involved in cellular processes, metabolic processes,response to stimulus, immune system, and so on. KEGG analysis results showed that the differentially expressed genes were mainly enriched in cellular processes, signal transduction, disease, metabolism pathways. This study found many genes and critical signaling pathways which affected the pathogenicity of ALV-J by transcriptome analysis and would provide new ideas for understanding the pathogenic mechanism of ALV-J at molecular level.

Key words: Wenshang Barred chicken, ALV-J, RNA-Seq, differentially expressed gene

中图分类号:

S831.2

张会永, 吴井生, 杨剑波, 俞燕, 薛倩, 殷建玫, 朱云芬, 朱静, 苏一军, 李国辉, 韩威. 基于RNA-Seq筛选ALV-J感染汶上芦花鸡肝差异表达致病相关基因[J]. 畜牧兽医学报, 2020, 51(8): 2003-2011.

ZHANG Huiyong, WU Jingsheng, YANG Jianbo, YU Yan, XUE Qian, YIN Jianmei, ZHU Yunfen, ZHU Jing, SU Yijun, LI Guohui, HAN Wei. Application of RNA-Seq Technology for Screening the Differentially Expressed Pathogenic Genes in Wenshang Barred Chicken Liver Infected by Subgroup J Avian Leukosis Virus[J]. Acta Veterinaria et Zootechnica Sinica, 2020, 51(8): 2003-2011.

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基于RNA-Seq筛选ALV-J感染汶上芦花鸡肝差异表达致病相关基因

Application of RNA-Seq Technology for Screening the Differentially Expressed Pathogenic Genes in Wenshang Barred Chicken Liver Infected by Subgroup J Avian Leukosis Virus

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