摘要
【目的】昆虫种群密度的升高往往成为一种胁迫因子。粘虫在低密度下形成散居型,而在高密度下则形成群居型。本研通过比较hsc70在两型粘虫中的表达模式,究旨在探索hsc70在粘虫Mythimna separata响应密度胁迫中的作用。【方法】采用RT-PCR和RACE技术克隆粘虫hsc70cDNA全长序列,利用生物信息学软件分析该基因及其编码蛋白质的序列特性;运用实时荧光定量PCR技术(qRT-PCR)分析该基因在两型粘虫不同发育阶段(卵、1-6龄幼虫、蛹和成虫)、5龄幼虫不同组织(头、表皮、前肠、中肠、后肠和马氏管)以及群居和分离处理后mRNA表达量的变化。【结果】克隆得到的HSC70基因命名为Mshsc70(GenBank登录号:MH669276),全长cDNA长2 184 bp,开放阅读框为1 965 bp,编码653个氨基酸。序列分析发现MsHSC70具有HSP70家族典型的结构域,即IDLGTTYS(11-18aa),IFDLGGGTFDVSLL (198-211 aa)和IVLVGGSTRIPKVQQ (341-355 aa)。序列比对结果显示,MsHSC70与斜纹夜蛾Spodopteralitura,棉铃虫Helicoverpaarmigera,大螟Sesamiainferens、甘蓝夜蛾Mamestra brassicae HSC70的相似性高达99%;基于HSC70的系统发育分析显示,粘虫与夜蛾科(Noctuidae)昆虫的亲缘关系最近。Mshsc70在两型粘虫的各个发育阶段均表达,hsc70的表达量在1龄和5龄中的表达量较散居型显著上调,而在2-3龄中的表达量显著下调。除表皮和前肠外,群居型粘虫的其他组织中hsc70的表达量显著高于散居型。散居型粘虫经群聚饲养36h后,hsc70显著上调表达,表达量是对照的3.17倍,而群居型粘虫经分离饲养36 h后,hsc70的表达量与对照无显著差异。【结论】粘虫的种群密度是决定粘虫hsc70表达的一个重要因子。
[Objectives] High population density causes stress to insect populations. Mythimna separata has solitary and gregarious phases that occur at low and high population densities, respectively. This study aims to explore the role of hsc70 under crowding stress by comparing the expression profiles of hsc70 in the two phases of the M. separata. [Methods] The complete cDNA sequence of hsc70 was cloned by RT-PCR and RACE. Bioinformatics programs were used to analyze the sequence characteristics of the gene and its amino acid sequences. The expression levels of hsc70 in different developmental stages(egg, 1 st-6 th instar larva, pupa and adult) and tissues of 5 th instar larvae(head, epidermis, fore-gut, mid-gut, hindgut and Malpighian tubules) from different phases(solitary and gregarious) were detected by Quantitative Real-time PCR(qRT-PCR). [Results] The complete cDNA of hsc70 obtained from M. separata(named as Mshsc70) was 2 184 bp in length(GenBank accession no.: MH669276), including a 1 965 bp opening reading frame(ORF) encoding 653 amino acids. Sequence analysis indicates that MsHSC70 has typical motifs of the HSP70 family, including IDLGTTYS(11-18 aa), IFDLGGGTFDVSLL(198-211 aa) and IVLVGGSTRIPKVQQ(341-355 aa). Phylogenetic analysis indicates that HSC70 s from Noctuid moths could be assigned to one well-supported cluster. Mshsc70 was expressed in all developmental stages of both phases, its expression in 1 st and 5 th instar larvae of gregarious individuals was significantly up-regulated compared to solitary individuals, whereas its expression in 2 nd and 3 rd instar larvae was obviously down-regulated. The differential expression of hsc70 in different tissues was higher in gregarious than in solitary individuals, except in the epidermis and foregut. hsc70 transcripts were significantly up-regulated(3.17 times of that the control) in solitary M. separata larvae after these were kept in crowded conditions for 36 h, but transcript abundance did not change in gregarious larvae that were kept isolated for 36 h. [Conclusion] Population density may be an important factor determining hsc70 expression in M. separata
作者
李鸿波
戴长庚
张昌容
何永福
LI Hong-Bo;DAI Chang-Geng;ZHANG Chang-Rong;HE Yong-Fu(Institute of Plant Protection,Guizhou Academy of Agricultural Sciences,Guiyang 550006,China)
出处
《应用昆虫学报》
CAS
CSCD
北大核心
2018年第5期825-833,共9页
Chinese Journal of Applied Entomology
基金
公益性(农业)科研专项(201403031)
贵州省自然科学基金(黔科合J字[2014]2121)
国家自然科学基金(31601633)
贵州省农科院博士启动项目(201301
企业服务行动计划(黔2015-4012)
