二化螟dsRNA降解酶基因的克隆与表达分析  

Cloning and expression analysis of double-stranded RNA degrading enzyme genes in the rice stem borer Chilo suppressalis

作　　者：彭英传 江婷 朱玉麟 张万娜 张晶 韩召军 肖海军[1,3] Peng Yingchuan;Jiang Ting;Zhu Yulin;Zhang Wanna;Zhang Jing;Han Zhaojun;Xiao Haijun(Institute of Entomology,Jiangxi Agricultural University,Nanchang 330045,Jiangxi Province,China;Key Laboratory of Integrated Management of Crop Diseases and Pests,Ministry of Education,College of Plant Protection,Nanjing Agricultural University,Nanjing 210095,Jiangsu Province,China;School of Grassland Science,Beijing Forestry University,Beijing 100083,China)

机构地区：[1]江西农业大学昆虫研究所,南昌330045 [2]南京农业大学植物保护学院,农作物生物灾害综合治理教育部重点实验室,南京210095 [3]北京林业大学草业与草原学院,北京100083

出　　处：《植物保护学报》2023年第3期610-619,共10页Journal of Plant Protection

基　　金：国家自然科学基金(32102223);江西省自然科学基金(20224BAB215019);江西省教育厅科学技术研究项目(GJJ180216)。

摘　　要：为解析二化螟Chilo suppressalis体内参与降解双链RNA(double-stranded RNA,dsRNA)的关键核酸酶的功能,克隆二化螟不同的非专一性核酸酶(non-specific nuclease,NUC)基因,并对这些基因进行生物信息学分析和组织定量表达分析,同时对dsRNA降解酶(dsRNA degrading nuclease,dsRNase)活力的组织分布进行研究。结果显示,共克隆获得5个NUC基因,其中有4个编码dsRNase亚家族基因(CsdsRNase1~CsdsRNase4)和1个编码Endonuclease G亚家族基因(CsEndoG)。5个NUC基因的开放阅读框核苷酸序列长度范围为828~1338 bp,编码275~445个氨基酸残基,其分子量大小为31.68~49.57 kD,预测等电点为5.48~9.42。CsdsRNase1和CsdsRNase2含有信号肽序列,两者相似度极高,且与家蚕Bombyx mori和斜纹夜蛾Spodoptera litura中具有dsRNA降解酶活力的dsRNase同源聚类;CsdsRNase1和CsdsRNase2均在二化螟中肠中高表达,这与中肠组织超高的dsRNA降解酶活力分布模式一致,表明这2个基因可能是中肠参与降解dsRNA的关键核酸基因。二化螟体内存在多条dsRNase基因序列,尤其是中肠中高表达的CsdsRNase1和CsdsRNase2可影响二化螟体内dsRNA的稳定性,从而影响RNA干扰效率。To elucidate the function of the key nucleases involved in the degradation of double-stranded RNA(dsRNA)in rice stem borer Chilo suppressalis,different non-specific nuclease(NUC)genes were cloned and subjected to bioinformatics analysis and tissue-specific expression analysis.The distribution of the activity of dsRNA degrading nucleases(dsRNase)in different tissues was also investigated.Five NUC genes were cloned,including four genes encoding the dsRNase subfamily(CsdsRNase1-Csd-sRNase4)and one gene encoding the endonuclease G subfamily(CsEndoG).The open reading frames of the five NUC genes ranged from 828 to 1338 bp,encoding 275 to 445 amino acid residues,with mo-lecular weights ranging from 31.68 to 49.57 kD and predicted isoelectric points ranging from 5.48 to 9.32.CsdsRNase1 and CsdsRNase2 contained putative signal peptide sequences,and they shared high sequence identity.CsdsRNase1 and CsdsRNase2 were clustered in the same clade with dsRNase en-zymes that showed dsRNA-degrading activity in Bombyx mori and Spodoptera litura.CsdsRNase1 and CsdsRNase2 were predominantly expressed in the midgut,consistent with the high distribution of dsRNA degrading activity in this tissue,indicating that these two genes might be key nucleases in-volved in dsRNA degradation in the midgut.The presence of multiple dsRNase gene sequences in C.suppressalis,especially CsdsRNase1 and CsdsRNase2,which were highly expressed in the midgut,might affect the stability of dsRNA,thereby affecting RNA interference efficiency.

关 键 词：二化螟 非专一性核酸酶 基因克隆 RNA干扰 dsRNA降解酶 

分 类 号：S435.112.1[农业科学—农业昆虫与害虫防治]