杨朔 刘少凯 赵少轩 朱宇辰 徐磊 李童云 刘晓光 安世恒 魏纪珍

摘要 ：棉鈴虫Helicoverpa armigera是一种重要的农业害虫，本文旨在研究小分子热激蛋白在其生长发育过程、抵御高温及Cry1Ac杀虫蛋白中的功能。利用PCR结合RACE技术克隆了棉铃虫sHSP22.0（small heat shock protein 22.0）基因，通过生物信息学软件分析了棉铃虫sHSP22.0基因序列，利用实时荧光定量qRT-PCR分析了该基因在棉铃虫不同生长发育阶段和组织中的表达模式;并分析了该基因受高温及Cry1Ac全长蛋白的诱导效应。获得了棉铃虫sHSP22.0（GenBank登录号： XP_021196802.1） 761 bp cDNA片段，其开放阅读框576 bp，编码191个氨基酸，具有小分子热激蛋白典型的α-晶体结构域（α-crystallin domain，ACD）。sHSP22.0在棉铃虫4龄和5龄期特异性表达，尤其在5龄幼虫的表皮、中肠和后肠内特异性表达。该小分子热激蛋白受温度和Cry1Ac全长蛋白的诱导后显著高表达。sHSP22.0不仅在暴食期及肠道内特异性表达，而且响应Cry1Ac全长蛋白的诱导，表明它在棉铃虫消化吸收及抵御外源物的活动中可能起到重要的作用。

关键词 ：棉铃虫; 小分子热激蛋白; sHSP22.0; 温度; Cry1Ac全长蛋白

中图分类号：

Q 966

文献标识码： A

DOI： 10.16688/j.zwbh.2020009

Molecular cloning and expression profiling of small heat shock protein 22.0 in

Helicoverpa armigera （Lepidoptera： Noctuidae）

YANG Shuo， LIU Shaokai， ZHAO Shaoxuan， ZHU Yuchen， XU Lei， LI Tongyun，

LIU Xiaoguang， AN Shiheng， WEI Jizhen*

（State Key Laboratory of Wheat and Maize Crop Science， College of Plant Protection， Henan Agricultural University， Zhengzhou 450002， China）

Abstract

Helicoverpa armigera is an important agricultural pest. This study aims to explore the functions of small heat shock protein 22.0 gene （sHSP22.0） in the growth and development of cotton bollworm， and its role in suffering the high temperature and Cry1Ac protein. The full-length cDNA sequence of sHSP22.0 gene was cloned from H.armigera by using PCR and RACE （rapid amplification of cDNA ends） technology， and the sHSP22.0 gene was analyzed by bioinformatics related molecular software. Real-time quantitative PCR （qRT-PCR） was used to assess the expression patterns of sHSP22.0 in different developmental stages and tissues of cotton bollworm. Meanwhile， the expression levels of sHSP22.0 were also checked in the larval after heat and full-length Cry1Ac protein treatments. The results showed that the full-length cDNA sequence of sHSP22.0 （GenBank accession no： XP_021196802.1） was 761 bp with an open reading frame （ORF） 576 bp in length， encoding 191 amino acids， and it had the typical α-crystallin domain （ACD）. The sHSP22.0 gene was specially expressed in 4th and 5th instar larvae， and especially expressed in midgut， hindgut and cuticle of 5th instar larvae. After the heat and full-length Cry1Ac protein treatments， sHSP22.0 showed special expression in 5th instar larvae. The sHSP22.0 gene specially expresses in the gluttony period and gut of H.armigera， and importantly， it is induced by full-length Cry1Ac protein. All the results indicate sHSP22.0 may participate in the activities of the digestion and absorption， and may have the function in defensing against exogenous substrates.

Key words

Helicoverpa armigera; small heat shock protein; sHSP220; temperature; full-length Cry1Ac protein

热激蛋白（heat shock proteins，HSPs）在生物体内广泛存在，以应激蛋白和分子伴侣的方式响应外界生物和非生物因子的压力，参与蛋白质的装配、折叠和跨膜传导等众多的生物过程[12]。按照序列的同源性及分子量的大小，热激蛋白常被分为4类：HSP90（85～90 kDa）、HSP70（68～73 kDa）、HSP60和小分子量sHSP家族[3]。在自然界中，昆虫个体较小又是变温动物，由于长期适应环境的变化，昆虫体内进化出了种类繁多的热激蛋白[4]。其中小分子热激蛋白（sHSP）的数目尤为繁多，它们具有保守的α-晶状体结构域，但相对于大分子的热激蛋白，其序列不保守且功能复杂。……