Evidence for gene duplication in the voltage-gated sodium channel gene of Aedes aegypti.
Evidence for gene duplication in the voltage-gated sodium channel gene of Aedes aegypti.
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DOI：
10.1093/emph/eot012
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发表时间：
2013-01
期刊：
影响因子：
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通讯作者：
Peixoto AA
中科院分区：
文献类型：
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作者：
Martins AJ;Brito LP;Linss JG;Rivas GB;Machado R;Bruno RV;Lima JB;Valle D;Peixoto AA
在这里，我们展示了埃及伊蚊NAV基因复制的第一个证据，这可能与杀虫剂抗性有关。复制的单倍型由一个带有特定突变的序列和另一个没有特定突变的序列组成，在自然种群中作为一种多态特征存在。背景和目的：电压门控钠通道基因(NAV)的突变称为kdr突变，在许多物种中与拟除虫菊酯和滴滴涕的抗药性有关。在蚊子登革热媒介埃及伊蚊中，除了kdr，其他多态允许将Aanav序列归类为A或B类型。在这里，我们指出了一系列证据，表明这些多态实际上与基因复制事件有关。方法：采用4种方法：(1)用等位基因特异性聚合酶链式反应(AS-PCR)对可能存在kdr突变的两个Aanav位点(Ile1011Met和Val1016Ile)进行基因分型；(2)部分Aanav基因的克隆和测序；(3)与特定谱系杂交并分析后代的基因类型；(4)用TaqMan实时定量聚合酶链式反应(TaqMan)进行拷贝数变异分析。结果：KDR基因1011和1016位点突变仅存在于A型序列中，而不存在于同一单倍型中。此外，虽然1011Met突变等位基因广泛分布，但没有检测到纯合子(1011Met/Met)。测序显示在一些个体中有三种不同的单倍型，提出了基因重复的假说，这一假说得到了特定杂交后代的基因频率的支持。此外，据估计，与敏感的参考菌株相比，选择抗药性的实验室菌株的钠通道基因拷贝数增加了5倍。结论和启示：这里发现的Aanav复制可能是最近对大量使用杀虫剂的适应性反应，在同一生物体中同时保持野生型和突变等位基因，赋予抗药性并减少其一些有害影响。
Herein, we show the first evidence of a duplication of the NaV gene of the mosquito Aedes aegypti, that might be involved in insecticide resistance. The duplicated haplotype is composed of one sequence with and another without a specific mutation, present in natural populations as a polymorphic trait. Background and objectives: Mutations in the voltage-gated sodium channel gene (NaV), known as kdr mutations, are associated with pyrethroid and DDT insecticide resistance in a number of species. In the mosquito dengue vector Aedes aegypti, besides kdr, other polymorphisms allowed grouping AaNaV sequences as type ‘A’ or ‘B’. Here, we point a series of evidences that these polymorphisms are actually involved in a gene duplication event. Methodology: Four series of methods were employed: (i) genotypying, with allele-specific PCR (AS-PCR), of two AaNaV sites that can harbor kdr mutations (Ile1011Met and Val1016Ile), (ii) cloning and sequencing of part of the AaNaV gene, (iii) crosses with specific lineages and analysis of the offspring genotypes and (iv) copy number variation assays, with TaqMan quantitative real-time PCR. Results: kdr mutations in 1011 and 1016 sites were present only in type ‘A’ sequences, but never in the same haplotype. In addition, although the 1011Met-mutant allele is widely disseminated, no homozygous (1011Met/Met) was detected. Sequencing revealed three distinct haplotypes in some individuals, raising the hypothesis of gene duplication, which was supported by the genotype frequencies in the offspring of specific crosses. Furthermore, it was estimated that a laboratory strain selected for insecticide resistance had 5-fold more copies of the sodium channel gene compared with a susceptible reference strain. Conclusions and implications: The AaNaV duplication here found might be a recent adaptive response to the intense use of insecticides, maintaining together wild-type and mutant alleles in the same organism, conferring resistance and reducing some of its deleterious effects.