草地贪夜蛾细胞外信号调节激酶基因克隆及对不同环境胁迫的响应

Cloning of extracellular signal regulated kinase gene of fall armyworm Spodoptera frugiperda and its expression under different environmental stresses

为探究草地贪夜蛾Spodoptera frugiperda细胞外信号调节蛋白激酶(extracellular signal regulated protein kinase,ERK)对外界环境胁迫的响应,采用逆转录聚合酶链反应(reverse transcriptionpolymerase chain reaction,RT-PCR)克隆草地贪夜蛾ERK基因,命名为SfERK,运用生物信息学方法分析其序列特征,采用实时荧光定量PCR(quantitative real-time PCR,RT-qPCR)技术分析SfERK基因在不同龄期、不同组织及A区段紫外辐射(ultraviolet-A radiation,UV-A)胁迫和低温、高温胁迫下的表达情况。结果表明,草地贪夜蛾SfERK(登录号:MZ061919)基因cDNA全长为2127 bp,开放阅读框为1212 bp,编码404个氨基酸,编码蛋白质相对分子量约44.48 k D,等电点为6.34。系统发育分析表明,SfERK与鳞翅目昆虫ERK基因亲缘关系最近。发育表达模式表明SfERK在卵期表达量最高;组织表达模式表明SfERK在成虫头部(去除触角和复眼)相对表达水平达最高。UV-A和低温胁迫均可诱导SfERK表达。UV-A胁迫下,雌雄成虫体内SfERK基因分别在不同处理时间下显著上调,分别在120 min和30 min时的表达量最大;4℃低温胁迫下,雌雄成虫体内SfERK基因呈先上升后下降的表达模式,分别在120 min和90 min时达到峰值;36℃高温胁迫下,草地贪夜蛾雌成虫体内SfERK基因在高温胁迫下均显著下调;雄成虫体内SfERK基因呈先下降后上升再下降趋势,在90 min时表达量最大。表明SfERK基因在草地贪夜蛾响应环境胁迫的分子机制中可能发挥着重要作用。

To investigate the response of extracellular signal regulated protein kinase(ERK)of fall ar-myworm Spodoptera frugiperda to environmental stress,the ERK gene was cloned from S.frugiperda by reverse transcription-polymerase chain reaction(RT-PCR).It was named SfERK and its sequence characteristics were analyzed using bioinformatics methods.The relative expression levels of SfERK during different developmental stages,in different tissues of adults,and under UV-A,low and high tem-perature stresses were detected by quantitative real-time PCR(RT-qPCR).The cDNA of SfERK consist-ed of 2127 bp with an open reading frame of 1212 bp that encoded 404 amino acids.The relative mo-lecular weight of its putative protein was 44.48 kD with an isoelectric point of 6.34.Phylogenetic analysis showed that SfERK shared a high homology with ERK genes from other species of Lepidoptera.Devel-opmental expression profiling revealed that the expression level of SfERK was highest at the egg stage,and the tissue expression profiling showed that the expression level of SfERK was highest in the head without antennae and compound eyes.UV-A and low temperature stresses up-regulated the expression of SfERK gene.Under UV-A stress,the highest expression level of SfERK was detected in females at 120 min,while male adults reached the maximum at 30 min.Under 4℃,the expression level of SfERK in both female and male adults increased at first and then decreased with increasing exposure time,and reached the peak at 120 min and 90 min,respectively.Under 36℃,the expression level of SfERK in fe-male was significantly down-regulated,while its expression level in male decreased at first,then in-creased,and decreased again with increasing exposure time,and reached the maximum at 90 min.The ex-pression of SfERK in adult females was significantly down-regulated under high temperature stress.These results suggested that SfERK might play an important role in the molecular response of S.frugiperda to en-vironmental stress.