›› 2015, Vol. 58 ›› Issue (5): 487-495.doi:

• 研究论文 • 上一篇    下一篇

褐飞虱磷脂酰肌醇3激酶p85α亚基基因 NlPIK3R1的克隆与功能分析

陆潮峰# , 郝培应# ,马艳, 朱家骏, 冯娅琳, 俞晓平*   

  1. (中国计量学院生命科学学院, 浙江省生物计量及检验检疫技术重点实验室, 杭州 310018)
  • 出版日期:2015-05-20 发布日期:2015-05-20
  • 作者简介:陆潮峰, 男, 1989年生, 浙江绍兴人, 硕士研究生, 研究方向为昆虫分子生物学与生物化学, E-mail: lcf19890107@163.com; 郝培应, 男, 1975年生, 安徽寿县人, 博士, 副教授, 研究方向为昆虫分子生物学与生物化学, E-mail: haopeiy@163.com

Molecular cloning and function analysis of phosphatidyl inositol 3 kinase p85α subunit gene NlPIK3R1 in the brown planthopper, Nilaparvata lugens (Hemiptera: Delphacidae)

LU Chao-Feng# , HAO Pei-Ying# , MA Yan, ZHU Jia-Jun, FENG Ya-Lin, YU Xiao-Ping*   

  1. (Zhejiang Provincial Key Laboratory of Biometrology and Inspection & Quarantine, College of Life Sciences, China Jiliang University, Hangzhou 310018, China)
  • Online:2015-05-20 Published:2015-05-20

摘要: 【目的】PI3K信号通路在生物体中发挥重要功能,涉及糖和脂质的代谢、细胞和组织生长以及生物个体寿命等。本研究旨在探明该通路中磷脂酰肌醇3激酶(PI3K)在褐飞虱Nilaparvata lugens中的功能。【方法】根据转录组提供的PI3K p85α核心序列信息,应用cDNA末端快速克隆的技术(RACE)获得了编码PI3K p85α的基因NlPIK3R1的全长cDNA(GenBank登录号为KP635379),并应用荧光定量PCR和通过给成虫喂食dsNlPIK3R1对NlPIK3R1进行RNA干扰(RNAi)分别对该基因的表达规律和功能进行了研究。【结果】荧光定量PCR测定结果表明,NlPIK3R1在褐飞虱若虫和雄成虫中表达量均较低,但在怀卵雌成虫中大量表达。RNAi结果表明,给褐飞虱成虫喂食0.1和0.5 μg/μL dsNlPIK3R1均导致NlPIK3R1的表达明显受到抑制,高浓度组的抑制效果尤为明显。喂食dsNlPIK3R1对褐飞虱成虫具有极显著致死效果,高浓度组的褐飞虱成虫在饲喂第7天时存活率仅为37.5%,相比于空白对照组(87.00%)和dsGFP对照组(76.67%)均达到了极显著差异(P<0.01)。对NlPIK3R1的RNAi导致褐飞虱成虫羽化率下降和体重变轻。【结论】本研究结果显示,NlPIK3R1基因对褐飞虱的生存、生长和发育具有重要作用,可以作为防治褐飞虱的潜在靶标。

关键词: 褐飞虱, 磷脂酰肌醇3激酶, p85&alpha, NlPIK3R1基因, RNA干扰

Abstract: 【Aim】 PI3K signaling pathway plays important roles in many organisms involving in carbohydrate and lipid metabolism, cell and tissue growth, life span, etc. This study aims to study the function of phosphatidylinositol 3-kinase in the rice brown planthopper, Nilaparvata lugens. 【Methods】 The full-length cDNA of NlPIK3R1 gene was cloned by rapid-amplification of cDNA ends (RACE) according to its gene sequence information available in N. lugens transcriptome, and this gene encodes the p85α subunit of PI3K (PI3K p85α), with the GenBank accession number of KP635379. Then the real-time quantitative PCR and RNAi by feeding dsNlPIK3R1 to N. lugens adults were conducted to explore the expression pattern and the function of NlPIK3R1, respectively. 【Results】 The real-time quantitative PCR results showed that the NlPIK3R1 gene was slightly expressed in nymphs and male adults of N. lugens, but strongly expressed in gravid female adults. RNAi results showed that the expression of NlPIK3R1 was inhibited in the two dsNlPIK3R1 treatment groups (fed with 0.1 and 0.5 μg/μL dsNlPIK3R1, respectively), especially in the high concentration treatment group. Feeding of dsNlPIK3R1 led to the death of N. lugens adults. The survival rate of N. lugens adults in the high concentration treatment group significantly declined by 37.5% at 7 d after treatment, and showed significant differences (P<0.01) from those in the blank control group (87%) and the dsGFP group (76.67%). RNAi also resulted in a reduced eclosion rate and lighter body weight. 【Conclusion】 The results suggest that the NlPIK3R1 gene is very important for the survival and growth of N. lugens, and may serve as a potential target for controlling the brown planthopper.

Key words: Nilaparvata lugens, phosphatidylinositol 3-kinase, p85α, NlPIK3R1, RNA interference