棉铃虫BR-C Z4基因的克隆及表达分析  

Cloning and expression analysis of BR-C Z4 gene in cotton bollworm Helicoverpa armigera

作　　者：刘静 张连军 胡得琴 张亚林 刘小宁 Liu Jing;Zhang Lianjun;Hu Deqin;Zhang Yalin;Liu Xiaoning(Xinjiang Key Laboratory of Biological Resources and Genetic Engineering,College of Life Science and Technology,Xinjiang University,Urumqi 830017,Xinjiang Uygur Autonomous Region,China)

机构地区：[1]新疆大学生命科学与技术学院,新疆生物资源基因工程重点实验室,乌鲁木齐830017

出　　处：《植物保护学报》2023年第4期962-971,共10页Journal of Plant Protection

基　　金：国家自然科学基金(31972279);新疆维吾尔自治区天山雪松计划(2017xs02)。

摘　　要：为明确转录因子广泛锌指复合物(broad-complex,BR-C)Z4亚型在棉铃虫Helicoverpa armigera生长发育过程中的作用,基于2-十三烷酮(2-tridecanone,2-TD)处理的棉铃虫幼虫中肠转录组数据克隆获得BR-C Z4的开放阅读框(open reading frame,ORF)序列,对其氨基酸序列和编码蛋白质进行生物信息学分析,并利用实时荧光定量PCR(real-time fluorescence quantitative PCR,qPCR)技术分析BR-C Z4基因在棉铃虫不同龄期、不同组织和2-TD处理6龄幼虫中的表达规律以及其和Ⅳ型几丁质酶(chitinase IV,CHT4)编码基因在4~6龄初、末期幼虫中的表达规律。结果表明,克隆获得的棉铃虫BR-C Z4基因的ORF为1347 bp,编码448个氨基酸,预测编码蛋白质的分子量和理论等电点分别为49.8 kD和7.75,且主要定位于细胞核中。系统进化分析结果显示,棉铃虫BR-C Z4与家蚕Bombyx mori和烟草天蛾Manduca sexta的BR-C Z4亲缘关系最近。BR-C Z4基因在棉铃虫整个生长期均有表达,在3龄期的相对表达量最低而在预蛹期的相对表达量最高;该基因在棉铃虫6龄幼虫的脂肪体、中肠、体壁和头部均有表达,在头部的相对表达量最低,在中肠中的相对表达量最高;BR-C Z4和CHT4基因在棉铃虫4~6龄末期幼虫中的相对表达量均显著高于4~6龄初期幼虫;相关性分析结果显示BR-C Z4和CHT4的相对表达量呈极显著正相关;随着2-TD处理浓度的增加,棉铃虫BR-C Z4基因的相对表达量总体呈现先上升后下降的趋势,其中15 mg/g浓度处理20 h时BR-C Z4基因相对表达量达到峰值,而20 mg/g浓度处理6 h时BR-C Z4基因相对表达量降到最低,为对照的22.73%。推测BR-C Z4基因在棉铃虫变态发育和应对2-TD胁迫过程中发挥着重要作用。To investigate the role of transcription factor broad-complex(BR-C)subtype Z4 in the growth and development of cotton bollworm Helicoverpa armigera,the open reading frame(ORF)sequence of BR-C Z4 was cloned from larval midgut based on the transcriptomic data,which was obtained from midgut of H.armigera treated with 2-tridecanone(2-TD),and its amino acid sequence and protein structure were analyzed with bioinformatics.Real-time fluorescence quantitative PCR(qPCR)was used to detect the expression profiles of BR-C Z4 gene in different developmental stages,different tissues and its expression level in the 6th instar larvae treated with 2-TD,and the expression profiles of BR-C Z4 and chitinase IV(CHT4)genes in early stages and late stages of 4th-6th instar larvae.The results showed that the ORF of BR-C Z4 gene in H.armigera was 1347 bp in length,encoding 448 amino acids.The predicted molecular weight and the isoelectric point of the BR-C Z4 protein were 49.8 kD and 7.75,and it was mainly located in the nucleus of H.armigera.Phylogenetic analysis showed that BR-C Z4 of H.armigera was closely related to BR-C Z4 of Bombyx mori and Manduca sexta.BR-C Z4 gene was expressed in all growth periods of H.armigera,and the expression level was lowest in the 3rd instar larvae and highest in the pre-pupa stage.BR-C Z4 gene was expressed in fat body,midgut,integument and head of 6th instar larvae of H.armigera,with the lowest expression level in head and the highest in midgut.The expression levels of BR-C Z4 gene and CHT4 gene from the late stage from 4th to 6th instar larvae were correspondingly higher than those from the early stage.There was a significant positive correlation between the expressions of BR-C Z4 and CHT4.The relative expression of BR-C Z4 gene of H.armigera treated with 2-TD increased firstly and then decreased,and the relative expression level of BR-C Z4 gene of H.armigera treated with 15 mg/g 2-TD for 20 h reached the peak.The relative expression level of BR-C Z4 gene decreased to 22.73%of the control group at 6 h

关 键 词：棉铃虫 广泛锌指复合物 克隆 时空表达 2-十三烷酮 

分 类 号：S435.622.3[农业科学—农业昆虫与害虫防治]