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氯离子通道与农业害虫的抗药性发生有密切关联。本研究结合转录组测序及荧光定量PCR技术,鉴定和分析黄曲条跳甲Phyllotreta striolata(Fabricius)谷氨酸门控氯离子通道(GluCl)的基因序列特征、功能及基因表达谱。结果表明:本研究所获得的GluCl cDNA序列长度为1430bp,开放阅读框(open reading frame,ORF)长为1344bp,编码447个氨基酸残基。其推测蛋白分子主要含神经递质分子胞外结合域和胞内跨膜域两大功能域,其亲水的胞外结合域又含有2个由半胱氨酸二硫键形成的氨基酸残基桥环,表现出GluClα亚基的典型特征。cDNA序列的系统发育分析表明,该GluCl与其他昆虫的GluClα高度同源,而在脊椎动物中则与γ-GABA或甘氨酸配体门控氯离子通道同源。荧光定量PCR分析表明,该GluCl基因在黄曲条跳甲雌雄成虫的不同部位中都有表达,但是在头部的表达量非常高,如雄虫头部的表达量是其精巢的65.7倍、是其中肠的227.5倍,揭示GluCl基因在中枢神经组织中具有重要作用。本研究为进一步研究GluCl介导的黄曲条跳甲抗药性的发生及其分子机理奠定了基础。
Chloride channels are closely related to the resistance of agricultural pests. In this study, transcriptome sequencing and fluorescence quantitative PCR were used to identify and analyze the gene sequence, function and gene expression profiles of glutamate-gated chloride channel (Phyllotreta striolata (Fabricius)). The results showed that the length of GluCl cDNA obtained in this study was 1430bp and the open reading frame (ORF) length was 1344bp, encoding 447 amino acid residues. It is presumed that the protein molecules mainly contain the two domains of neurotransmitter molecule extracellular binding domain and intracellular transmembrane domain, and the hydrophilic extracellular binding domain contains two amino acid residues formed by cysteine disulfide bond Bridged ring, showing typical features of GluCl alpha subunit. Phylogenetic analysis of the cDNA sequence showed that this GluCl is highly homologous to GluClα of other insects, whereas in vertebrates it is homologous to γ-GABA or glycine-ligand-gated chloride channels. Quantitative real-time PCR analysis showed that the GluCl gene was expressed in different parts of both adult and male Bemisia tabaci, but its expression level in the head was very high. For example, the expression level of the GluCl gene was 65.7 times that of the testis, 227.5 times that of the midgut, revealing that the GluCl gene plays an important role in central nervous system. This study laid the foundation for the further study on the occurrence and molecular mechanism of GluCl-mediated resistance of Bentulina thaliana.