中华蜜蜂Takeout(AcTO1)基因的克隆和表达分析  

Cloning and expressional analysis of a takeout gene in Apis cerana

作　　者：陈艺杰 解幸承 吴双[1,2,4] 吴珍红[1,2] 李显春[4,5] 缪晓青[1,2,3] 

机构地区：[1]福建农林大学蜂学学院 [2]福建农林大学蜂疗研究所 [3]天然生物毒素国家地方联合工程实验室,福州350002 [4]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 [5]Department of Entomology and BIO5 Institute,The University of Arizona,Tucson,AZ 85721,USA

出　　处：《中国蜂业》2017年第7期16-20,共5页Apiculture of China

基　　金：国家蜂产业技术体系建设专项资金(CARS-45-KXJ19)

摘　　要：Takeou(tTO)是一种昆虫节律性调控输出基因,广泛分布于涉及到化学感受和营养功能的相关组织。TO基因主要参与昆虫的生长发育、行为调节及多种生理代谢过程。为探究TO基因是否参与社会性昆虫的劳动分工,我们克隆并分析了中华蜜蜂ApisceranaTO基因的编码框序列,测定了该基因在哺育蜂和采集蜂各个组织部位的mRNA表达水平,旨在为深入研究该基因的功能提供参考。本研究通过RT-PCR的方法首次克隆获得AcTO1基因的cDNA序列,利用生物信息学软件分析了该基因的核苷酸序列和氨基酸序列,并利用荧光定量PCR(qPCR)技术检测了该基因在中华蜜蜂3种采集蜂(18日龄正常采集蜂、22日龄正常采集蜂和7日龄提前采集蜂)和2种哺育蜂(7日龄正常哺育蜂和22日龄超龄哺育蜂)头部、胸部和腹部的表达量。本研究克隆获得的中华蜜蜂TO(命名为AcTO1)基因的cDNA序列长度为1058bp,开放阅读框(ORF)长度为738bp,编码246个氨基酸,预测蛋白分子量为27.09kDa,理论等电点为8.40。AcTO1蛋白的信号肽位于1-17位氨基酸,无跨膜结构,属于分泌型蛋白。AcTO1的氨基酸序列种含有一个JHBP超家族保守结构域。qPCR结果显示,AcTO1在3种采集蜂和2种哺育蜂的各组织均有表达,其中表达量由高到低依次为头部、胸部、腹部。AcTO1基因的表达具有组织依赖型,主要表达于头部,且所有采集蜂的头部表达量均高于哺育蜂,由此说明该基因可能参与蜜蜂的劳动分工。本研究克隆了中华蜜蜂takeout基因的全长cDNA序列,并分析了该基因的序列特征和表达谱,结果表明该基因可能参与调控蜜蜂的劳动分工。The takeout （TO） gene is a circadian clock-regulated output gene abundantly expressed in several tissues that are related to perception of chemical signals and nutrition acquisition in insects. TO protein is known to be involved in growth and development, behavior regulation and metabolism of the solitary insects. But it remains unclear whether TO protein is also involved in the division of labor in social insects. In order to address this general question and reveal its additional functions, we cloned the Apis cerana TO （AcTO1） gene and analyzed its expression levels in different tissues and developmental stages between nurse bees and forager bees. We obtained the eDNA sequence of AcTO1 by RT-PCR and analyzed its nucleotide and amino acid sequences using bioinformatics software. We also measured its expression levels in the head, thorax and abdomen of three types of forager bees （18-day old normal foragers, 22-day old normal foragers, 7-day old Young/precocious foragers） and two types of nurse bees （7-day old normal young nurses and 22-day old over-aged nurses） by veal-time quantitative PCR （qRT-PCR）. The AcTO1 cDNAsequence obtained by reverse transcriptase polymerase chain reading frame （ORF） of 738 bp, encoding a protein of 246 reaction （RT-PCR） was 1058 bp in length. It has an open amino acids. The predicted molecular weight of ACTO1 was 27.09 kDa and its theoretical isoelectric point was 8.40. ACTO1 protein contains a N-terminal signal peptide of 17 amino acids but lacks transmembrane domains, suggesting that ACTO1 was a secreted protein. ACTO1 protein also has an highly conserved JHBP （juvenile hormone binding protein） domain. In the three kinds of foragers and two kinds of nurses, ACTO1 expressed at the highest level in the head, followed by the thorax, and then the abdomen. Moreover, ACTO1 transcripts in the head were significantly higher in all the three kinds of forager bees than in the two kinds of nurse bees. Such expression profile implies that ACTO1 was probably assoc

关 键 词：蜜蜂 劳动分工 takeout基因 基因克隆 

分 类 号：S891[农业科学—特种经济动物饲养]