番茄潜叶蛾CYP9A304v2基因的克隆、时空表达及对乙基多杀菌素的表达响应
Cloning, spatiotemporal expression and response to spinetoram, of the Tuta absoluta CYP9A304v2 gene
王圣宇1** 解津刚1** 颜国荣2 林明灏1 王鑫海1 王圣斐1 王建民1 李玉珊2 刘小宁1***
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DOI：10.7679/j.issn.2095-1353.2026.075
作者单位：1. 新疆大学生命科学与技术学院，新疆生物资源基因工程重点实验室，乌鲁木齐 830017；2. 新疆维吾尔自治区农业科学院作物研究所，乌鲁木齐 830091
中文关键词：番茄潜叶蛾；CYP9A304v2；乙基多杀菌素；时空表达；生物信息学；分子对接
英文关键词：Tuta absoluta; CYP9A304v2; spinetoram; spatiotemporal expression; bioinformatics; molecular docking
中文摘要：【目的】 世界性入侵害虫番茄潜叶蛾Tuta absoluta对生物杀虫剂乙基多杀菌素表现出逐渐增强的抗药性，本研究旨在探究CYP9亚家族基因CYP9A304v2在番茄潜叶蛾生长发育及代谢乙基多杀菌素过程中的潜在功能。【方法】 本研究通过基因克隆、生物信息学分析、时空表达谱检测、胁迫诱导实验及分子对接等技术，对CYP9A304v2基因的序列特征、系统发育关系、在不同发育阶段以及乙基多杀菌素胁迫条件下的表达模式、以及其蛋白与药剂的结合能力进行分析。【结果】 CYP9A304v2基因开放阅读框长度为1 596 bp，编码了531个氨基酸，系统发育分析揭示番茄潜叶蛾CYP9A304v2属于CYP6家族。时空表达谱显示该基因在番茄潜叶蛾幼虫4龄和蛹期特异性高表达，且在乙基多杀菌素胁迫下显著上调表达（P ＜ 0.05）。分子对接结果显示，CYP9A304v2蛋白与乙基多杀菌素具有较强的结合亲和力，结合自由能均低于﹣37.7 kJ/mol。【结论】 时空表达谱显示CYP9A304v2在番茄潜叶蛾幼虫4龄及蛹期特异性高表达，且其蛋白与乙基多杀菌素具有强结合亲和力，暗示该基因可能在幼虫-蛹转变期间及对乙基多杀菌素的解毒代谢起关键作用。本研究为番茄潜叶蛾抗药性机制研究提供了候选基因及理论依据。
英文摘要：[Aim] To investigate the potential role of the CYP9 subfamily gene CYP9A304v2 in the development of Tuta absoluta and the response of this gene to spinetoram, a biopesticide to which T. absoluta is becoming increasingly resistant. [Methods] Gene cloning, bioinformatics analysis, spatiotemporal expression profiling, stress induction experiments, and molecular docking were employed to characterize the sequence features and phylogenetic relationships of CYP9A304v2. In addition, expression patterns in different developmental stages and in response to spinetoram were measured, together with an evaluation of the binding affinity between the CYP9A304v2 protein and spinetoram. [Results] The open reading frame (ORF) of CYP9A304v2 was 1 596 bp in length, and encoded 531 amino acids. Phylogenetic analysis revealed that T. absoluta CYP9A304v2 belongs to the CYP6 family. Spatiotemporal expression profiling demonstrated specific high expression of CYP9A304v2 in the 4th instar larval and pupal stages, and that its expression was significantly upregulated under spinetoram stress (P < 0.05). Molecular docking analyses indicates strong binding affinity between the CYP9A304v2 protein and spinetoram, with binding free energies consistently below -9 kcal/mol. [Conclusion] The high expression of CYP9A304v2 during the 4th instar larval and pupal stages, combined with its strong binding affinity for spinetoram, suggests that this gene may play a critical role in both the larval-pupal transition and the detoxification of spinetoram. These results identify a candidate gene that could be targeted to control T. absoluta, and provide new information on insecticide resistance mechanisms in this species.