摘要
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 channel was closely related to insect resistance to pesticides. ESTs of glutamate mediated chloride channel （GluCl） gene were clustered and identified from transcriptome data of Phyllotreta striolata （Fabricius）. The expression of GluCl gene in different tissues of P. striolata adults was profiled by quantitative real-time PCR. The results indicated that the cDNA of this gene is 1 430 bp in length with an open reading frame （ORF） encoding 447 amino acids. The putative protein molecule contains two functional domains： a neurotransmitter-gate ion-channel ligation binding domain and a neurotransmitter-gate ion-channel transmembrane domain. There are two cysteine loops in ligation binding domain regions demonstrating a typical characteristic of GluClα subunit. Phylogenetic analysis of the complete cDNA sequence showed that the GluCl of P. striolata is highly homologous to GluClα of other insects. Quantitative real-time PCR assay showed that a relatively higher expression amount of GluCl gene was found in head than in other organs including antenna, midgut, testis, ovary, fore legs, middle legs and hind legs. The expression level of GluCl gene in head of male adult was 65.7 times as high as that in testis and 227.5 times in midgut, suggesting its important mediation in center nervous system. This study provides useful information for studying molecular mechanism of pesticide resistance mediated by GluCl of P. striolata.
出处
《昆虫学报》
CAS
CSCD
北大核心
2012年第2期233-240,共8页
Acta Entomologica Sinica
基金
国家自然科学基金项目(31101500)
关键词
黄曲条跳甲
抗药性
谷氨酸门控氯离子通道
Α亚基
荧光定量PCR
Phyllotreta striolata
pesticide resistance
glutamate-gated chloride channel （GluCl）
α subunit
real-time PCR
