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【目的】基于烟粉虱Bemisia tabaci转录组数据,系统分析了烟粉虱解毒代谢酶基因在噻虫嗪抗性品系中的表达模式,探讨了这些基因在烟粉虱不同发育阶段差异表达的生物学意义。【方法】分别收集室内长期饲养的烟粉虱噻虫嗪抗性和敏感品系的卵、4龄若虫和刚羽化1 d的雌成虫,在烟粉虱转录组数据库中挑选8 394条解毒代谢相关基因设计探针,通过探针杂交,得到烟粉虱噻虫嗪抗性品系表达谱芯片,比较了这些基因在抗性烟粉虱3个不同发育阶段的表达情况。并随机挑选了9个基因,在抗感品系间3个不同发育阶段进行了荧光定量PCR验证。【结果】在抗性烟粉虱的卵和4龄若虫发育阶段,共有3 424个差异表达基因,其中489个是编码3类解毒代谢酶(羧酸酯酶、谷胱甘肽-S-转移酶和细胞色素P450多功能氧化酶)的基因;有14个基因在4龄若虫发育阶段过量表达,其中10个为P450基因,4个为GST家族基因。在抗性烟粉虱的4龄若虫和雌成虫发育阶段,总共有1 273个差异表达基因,193个为3类解毒代谢酶家族的基因,其中有9个P450家族基因在雌成虫期的表达量超过4龄若虫期的10倍。此外,表达谱芯片分析还筛选到了一些候选抗性基因。qRT-PCR验证显示在这些候选基因中,与敏感品系相比,9个基因在抗性烟粉虱的3个不同发育阶段表达上调,其中GST基因家族的p_06027和P450基因p_06013在抗性品系的卵和4龄若虫中过量表达;p_05885和p_07806和编码CYP6家族蛋白的p_00988在抗性品系的4龄若虫期的表达量上调;p_05916和p_00478在抗性品系卵和4龄若虫期表达量很低,而在成虫期过量表达;而p_00059和p_00428在抗性品系雌成虫发育阶段表达量显著上调,其中编码CYP4C1的p_00059的差异表达倍数在雌成虫期约为15.15倍。【结论】表达谱芯片分析结果提示,CYP6和CYP4C1基因的过量表达可能会是烟粉虱抗性产生的机制之一。解毒代谢酶基因在烟粉虱不同发育阶段的特异性表达,可能与其在抵御杀虫剂胁迫时体内能量的分布及有效利用率有关,也可能是害虫在环境选择压下的一种适应机制。
【Objective】 Based on the Bemisia tabaci transcriptome data of Bemisia tabaci, the expression patterns of the detoxification metabolizing enzyme genes of B. tabaci in the thiamethoxam resistance-resistant strains were systematically analyzed, and the biological characteristics of these genes in differentially expressed genes of B. tabaci at different developmental stages Significance of learning. 【Method】 The eggs, 4th instar nymphs and females newly emerging from thiamethoxam sensitive and susceptible lines of B. tabaci were collected from long-term indoors respectively. 8 394 detoxification and metabolism were selected in the transcriptome database of B. tabaci. Gene-designed probes were obtained by probe hybridization to get the thiamethoxam resistance strain expression profile chip of Bemisia tabaci, and the expression of these genes in three different developmental stages of B. tabaci was compared. Nine genes were randomly selected and verified by real-time PCR at three different developmental stages. 【Result】 A total of 3 424 differentially expressed genes were found in the eggs and 4th instar nymphs of B. tabaci, of which 489 genes encoding three kinds of detoxifying metabolizing enzymes (carboxylesterase, glutathione-S-transferase 14 genes were overexpressed at the 4th instar nymph development stage, of which 10 were P450 genes and 4 were GST family genes. In the 4th instar nymphs and female adults of B. tabaci, there were a total of 1 273 differentially expressed genes and 193 genes of the 3 detoxification metabolic enzymes. Among them, 9 P450 genes were expressed in female adults 10 times more than the 4th instar nymphs. In addition, cDNA microarray analysis also screened a number of candidate resistance genes. qRT-PCR validation showed that among these candidate genes, 9 genes were up-regulated at three different developmental stages of the resistant whiteflies, compared with the susceptible lines, in which the p_06027 and P450 genes of the GST gene family, p_06013, Eggs and 4th instar nymphs. The expression of p_05885 and p_07806 and the p_00988 encoding CYP6 family proteins were up-regulated in the 4th instar nymphs of the resistant lines. The expression levels of p_05916 and p_00478 in the resistant and 4th instar nymphs were low , And overexpression in adult stage. However, p_00059 and p_00428 were up-regulated in the developmental stages of female adults of the resistant strain, of which p_00059 encoding CYP4C1 was about 15.15 times more differentially expressed in female adults. 【Conclusion】 The results of expression profile microarray analysis suggest that overexpression of CYP6 and CYP4C1 genes may be one of the mechanisms of resistance to B. tabaci. The specific expression of detoxifying metabolic enzyme genes in different developmental stages of Bemisia tabaci may be related to the distribution and availability of energy in the body to resist insecticide stress and may also be an adaptation mechanism of pest insensitivity to environmental selection.