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【目的】保幼激素受体Methoprene-tolerant(Met)控制保幼激素信号传导途径中重要下游转录因子Krüppelhomolog 1(Kr-h1)的表达,调控昆虫的变态发育。本研究旨在探究褐飞虱Nilaparvata lugens Met基因与Kr-h1在褐飞虱变态发育中的功能。【方法】利用PCR技术扩增Nl Met基因的ORF序列。通过RNA干扰技术分别或同时沉默褐飞虱若虫的Nl Met和Nl Kr-h1基因,进一步研究其功能。【结果】克隆得到Nl Met,其开放阅读框包含1 185 bp,编码395个氨基酸,包括b HLH,PAS-A,PAS-B和PAC 4个结构域;其中PAS-B和PAC保守性较高,而b HLH和PAS-A保守性相对较低。采用RNA干扰技术沉默Nl Met和Nl Kr-h1基因后发现,单独对4龄若虫Nl Kr-h1基因干扰后,若虫阶段及初羽化雌虫和雄虫死亡率均显著上升(P<0.05);对5龄若虫Nl Kr-h1基因干扰后只有若虫死亡率上升(P<0.05),单独对Nl Met基因干扰后死亡率没有显著变化(P>0.05);二者共同沉默后的死亡率同单独对Nl Kr-h1沉默时类似。此外,我们还发现单独对4龄若虫Nl Kr-h1干扰后在雌虫中出现了生殖器畸形,虽然Nl Met干扰并未出现生殖器发育畸形现象,但二者共同干扰后,在若虫、初羽化雌雄成虫中畸形比例均显著增加(P<0.05)。【结论】Nl Met与下游转录因子Nl Kr-h1对褐飞虱变态发育起到重要作用,影响若虫到成虫的变态和外生殖器的发育。本研究有助于揭示Nl Met和Nl Kr-h1在昆虫变态发育中的重要作用。
【Objective】 Methoprene-tolerant (Met) regulates the expression of Krüppelhomolog 1 (Kr-h1), an important downstream transcription factor in juvenile hormone signaling pathways, and regulates insect metamorphosis. The aim of this study was to investigate the function of Nilaparvata lugens Met gene and Kr-h1 in the brown planthopper metamorphosis. 【Method】 The ORF of Nl Met gene was amplified by PCR. The functions of Nl Met and Nl Kr-hl genes of BPH nymphs were silenced separately or simultaneously by RNAi technology. 【Result】 Nl Met was cloned and contained 1 185 bp open reading frame (ORF) encoding 395 amino acids including 4 domains of b HLH, PAS-A, PAS-B and PAC. Among them, PAS-B and PAC were highly conserved , While b HLH and PAS-A are relatively less conserved. After silencing Nl Met and Nl Kr-h1 by RNA interference, the mortality of nymphs and virgins at the nymphal stages were significantly increased (P <0.05) after being interfered with the Nl Kr-h1 gene of 4th instar nymph respectively. Only the nymphal necrosis rate increased after the 5th instar nymphs (P <0.05), while the Nl Met gene alone had no significant difference (P> 0.05). The co-silencing mortality was the same as that of the control Nl Kr-h1 similar to silence. In addition, we found that genital abnormalities occurred in females after being interfered with the fourth instar nymphs, Nl Kr-h1. Although Nl Met did not show genital malformations, both of them interfered with each other in nymphs, The proportion of adult deformities were significantly increased (P <0.05). 【Conclusion】 Nl Met and its downstream transcription factor Nl Kr-h1 play an important role in the brown planthopper metamorphosis, affecting the nymphs and adult metamorphosis and the development of the external genitalia. This study will help reveal the important role of Nl Met and Nl Kr-h1 in insect metamorphosis.