猪ISG 20基因生物信息学分析及其抗PEDV感染的初步研究  

Bioinformatics Analysis of Porcine ISG 20 Gene and Preliminary Research of Its Effect against PEDV Infection

作　　者：王梦璐 张志杰 李之晗 聂晶晶 瞿云芝 张春红 刘志成 沈海燕 张建峰 孙亚妮[2] 李玉谷[1] WANG Meng-lu;ZHANG Zhi-jie;LI Zhi-han;NIE Jing-jing;QU Yun-zhi;ZHANG Chun-hong;LIU Zhi-cheng;SHEN Hai-yan;ZHANG Jian-feng;SUN Ya-ni;LI Yu-gu(College of Veterinary Medicine,South China Agricultural University,Guangzhou,Guangdong 510642,China;College of Veterinary Medicine,Northwest A&F University,Yangling,Shaanxi 712100,China;Institute of Animal Health,Guangdong Academy of Agricultural Sciences,Guangdong Key Laboratory for Livestock and Poultry Disease Control and Prevention,Guangdong Scientific Observation and Experimental Station for Veterinary Drugs and Diagnostic Technology,Ministry of Agriculture and Rural Affairs,Guangzhou,Guangdong 510640,China;Lingnan Modern Agricultural Science and Technology Guangdong Provincial Laboratory Maoming Branch Center,Maoming,Guangdong 525032,China)

机构地区：[1]华南农业大学兽医学院,广东广州510642 [2]西北农林科技大学动物医学院,陕西杨陵712100 [3]广东省农业科学院动物卫生研究所/广东省畜禽疫病防治研究重点实验室,广东广州510640 [4]岭南现代农业科学与技术广东省实验室茂名分中心,广东茂名525032

出　　处：《动物医学进展》2024年第10期7-14,共8页Progress In Veterinary Medicine

基　　金：猪禽种业全国重点实验室项目(2023QZ-NK13,ZQQZ-55);广东省农业农村厅"十四五"广东省农业科技创新十大主攻方向"揭榜挂帅"项目(2022SDZG02);广东省科学技术厅基础研究基金项目(2021A1515011125,2023A1515011681);广东省科学技术厅畜禽疫病防治研究重点实验室项目(2023B1212060040);岭南现代农业科学与技术广东省实验室茂名分中心自主科研项目(2022KF010);国家自然科学基金项目(31302101)。

摘　　要：为了研究猪干扰素刺激基因20(interferon-stimulated gene 20,ISG20)的序列信息及其表达情况,并探讨其对猪流行性腹泻病毒(Porcine epidemic diarrhea virus,PEDV)增殖的影响,以提取的猪小肠上皮细胞IPEC-J2总RNA为模板扩增猪ISG 20基因的CDS区序列,并应用生物信息学进行分析,进而构建pcDNA3.1-3×Flag-ISG20重组质粒转染入IPEC-J2细胞鉴定表达情况。此外,在转染重组质粒的细胞中感染PEDV分析ISG20对PEDV增殖的影响。结果表明,猪ISG 20基因CDS区的大小为516 bp,编码171个氨基酸,理论等电点(pI)为8.84;猪ISG20存在2个N-糖基化位点和23个磷酸化位点,主要定位于细胞骨架和细胞质中,以α-螺旋和无规则卷曲构成;构建的pcDNA3.1-3×Flag-ISG20重组质粒能够在IPEC-J2细胞中表达,蛋白大小约为19.6 ku,而且可以明显抑制PEDV的增殖;克隆了猪ISG 20基因,并且在IPEC-J2细胞中重组质粒pcDNA3.1-3×Flag-ISG20能够表达且发挥抑制PEDV增殖的作用,研究结果为深入探讨PEDV与ISG20的相互作用,以及以此为靶点筛选抗PEDV生物制剂提供了思路。This article aims to study the sequence information and expression of the porcine interferon stimulated gene 20(ISG20),and to explore its impact on the proliferation of porcine epizootic diarrhea virus(PEDV),using the extracted total RNA of porcine small intestine epithelial cells IPEC-J2 as a template to amplify the CDS region sequence of porcine ISG20 gene,and applying bioinformatics analysis to construct pcDNA3.1-3×Flag-ISG20 recombinant plasmid,the expression was identified by transfection into IPEC-J2 cells.In addition,the effect of ISG20 on PEDV proliferation in cells was analyzed after transfection with recombinant plasmids.The results showed that the size of the CDS region of the porcine ISG20 gene was 516bp,encoding 171 amino acids,and the theoretical isoelectric point(pI)was 8.84;Porcine ISG20 has 2 N-glycosylation sites and 23 phosphorylation sites,mainly located in the cytoskeleton and cytoplasm,which composed ofα-spirals and irregular curls.The pcDNA3.1-3×Flag-ISG20 recombinant plasmid can be expressed in IPEC-J2 cells,with a protein size of approximately 19.6 ku,and can significantly inhibit the proliferation of PEDV.In summary,the results of this study cloned the porcine ISG20 gene,and recombinant plasmid pcDNA 3.1-3 in IPEC-J2 cells×Flag-ISG20 can be expressed and exert an inhibitory effect on PEDV proliferation.This study provides insights into the interaction between PEDV and ISG20,as well as the screening of anti PEDV biologics using this as a target.

关 键 词：干扰素刺激基因20 生物信息学分析 真核表达 猪流行性腹泻病毒 

分 类 号：S852.651[农业科学—基础兽医学] S858.28[农业科学—兽医学]