粘虫脂动激素基因MsepAKH1的克隆与成虫期表达模式分析  

Molecular cloning and expression analysis of adipokinetic hormone gene in the adult oriental armyworm,Mythimna separata(Walker)

作　　者：解幸承 张蕾[2] 程云霞[2] 罗礼智[2] 仵均祥 江幸福[2] Xie Xingcheng;Zhang Lei;Cheng Yunxia;Luo Lizhi;Wu Junxiang;Jiang Xingfu(State Key Laboratory of Crop Stress Biology in Arid Areas, Northwest A＆F University, Yangling 712100, Shaanxi Province, China;State Key Laboratory for Biology of Plant Diseases and Insect Pests, Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beij ing 100193, China)

机构地区：[1]西北农林科技大学旱区作物逆境生物学国家重点实验室,陕西杨凌712100 [2]中国农业科学院植物保护研究所植物病虫害生物学国家重点实验室,北京100193

出　　处：《植物保护学报》2018年第3期552-560,共9页Journal of Plant Protection

基　　金：国家公益性行业(农业)科研专项(201403031);国家自然科学基金(31371947);北京市自然科学基金(6142017)

摘　　要：为明确脂动激素基因AKH在粘虫Mythimna separata(Walker)能源代谢过程中的作用,采用RT-PCR和RACE技术克隆得到粘虫脂动激素基因MsepAKH1的cDNA全长,并通过实时荧光定量PCR技术测定该基因在成虫期的组织与发育阶段表达模式。结果表明,粘虫脂动激素基因MsepAKH1的cDNA序列全长为449 bp,GenBank登录号为KP979739.1,开放阅读框为207 bp,编码68个氨基酸,具有昆虫脂动激素基因家族典型的结构特征;预测该基因编码的MsepAKH1蛋白分子量为7.61 kD,等电点为8.46,MsepAKH1的氨基酸序列与烟草天蛾Manduca sexta、二化螟Chilo suppressalis、小菜蛾Plutella xylostella、棉铃虫Helicoverpa armigera、家蚕Bombyx mori的AKH氨基酸序列同源性较高。MsepAKH1基因在粘虫初羽化雌蛾不同组织间的表达量差异显著,主要在头部和飞行肌内表达,分别约为同日龄雌蛾脂肪体内表达量的15.4倍与8.1倍,在脂肪体、卵巢与中肠内的表达量显著降低。对不同日龄雌蛾头部和飞行肌内的表达量检测发现,在7日龄雌蛾头部和3日龄雌蛾飞行肌内的表达量分别显著高于其它日龄雌蛾相同组织中的表达量,分别约为初羽化雌蛾相同组织表达量的2.4倍与1.6倍。表明MsepAKH1基因的表达因粘虫雌蛾组织与日龄间的不同而呈现动态变化。In order to clarify the biological function of the adipokinetic hormone gene（AKH） in the energy metabolism of oriental armyworm Mythimna separata（Walker）, the full-length cDNA sequence of AKH from M. separata was cloned by the reverse transcription PCR（RT-PCR） and rapid-amplification of cDNA ends（RACE）, which was named as MsepAKH1. The expression pattern of MsepAKH1 was analyzed with real time quantitative PCR（qPCR） method. The cDNA of MsepAKH1 was 449 bp（GenBank accession No. KP979739.1）, which included an open reading frame of 207 bp, encoding 68 amino acid residues, with the typical structural characteristics of the known insect AKH family. The predicted molecular mass and theoretical isoelectric point of MsepAKH1 was 7.61 kD and 8.46, respectively. Phylogenetic analysis indicated that the amino acid sequence of MsepAKH1 had a close relationship to the corresponding proteins from Manduca sexta, Chilo suppressalis, Plutella xylostella, Helicoverpa armigera and Bombyx mori. The real time quantitative PCR analysis showed that the relative expression levels of MsepAKH1 were significantly different in various tissues of one day-old female adults. MsepAKH1 mostly expressed in the head and flight muscle, which accounted for nearly 15.4 and 8.1 times higher than that in the fat body of the newly emerged female adults, respectively. But it very lowly expressed in the fat body, ovary and midgut of the newly emerged female adults. Compared with that in the different developmental stages, the relative expression levels of MsepAKH1 in the head of the seven day-old and the flight muscle of the three day-old female adults were significantly higher than those in the same tissues of other day-old female adults, which accounted for nearly 2.4 and 1.6 times higher than that of the newly emerged female adults, respectively. In conclusion, the expression of MsepAKH1 varied with the tissues and day-old of the female adults.

关 键 词：粘虫 脂动激素基因 克隆 表达模式 

分 类 号：S433.4[农业科学—农业昆虫与害虫防治]