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| 利用RNAi分析松材线虫ncr基因功能 | |
| 引用本文： | 王殿东,陈国华,潘丽梅.利用RNAi分析松材线虫ncr基因功能[J].植物病理学报,2018,48(4):518-526. |
| 作者姓名： | 王殿东 陈国华 潘丽梅 |
| 作者单位： | 长江师范学院,重庆 408100; 中国农业科学院蔬菜花卉研究所,北京 100081 |
| 基金项目： | 国家自然科学基金(31300309);重庆市基础科学与前言技术研究专项资助(cstc2014jcyjA00048);长江师范学院校级项目(2014QNRC04、2015XJXM06) |
| 摘 要： | 由松材线虫(Bursaphelenchus xylophilus)侵染引起的松材线虫病是松属植物的重要病害之一,其繁殖和传播速度快,防治困难。RNAi技术是目前研究植物寄生线虫基因功能的重要手段。本文利用农杆菌介导灰葡萄孢菌表达松材线虫ncr基因的dsRNA(PDH-RH-NCR),并连续多代喂饲松材线虫使ncr基因表达稳定下调后分析其表型。结果表明,在转有PDH-RH-NCR的灰葡萄孢菌上连续培养10代的松材线虫,ncr基因表达量显著下调13倍。ncr基因表达下调后,线虫发育缓慢,繁殖力也显著降低;处理4 d后发育成成虫的百分率为61.75%,显著低于PDH-RH-s GFP(89.75%)和空载体(88.75%)处理;而9 d的繁殖量则较PDH-RH-s GFP和空载体处理分别少2 314和2 347条。松材线虫ncr基因表达下调后,4龄幼虫平均寿命(20.9 d)和体内脂肪量(1.340 nmol)则显著高于PDH-RH-s GFP(15.5 d,0.336 nmol)和空载体(15.2 d,0.323 nmol)处理。通过喂饲法RNAi发现松材线虫ncr基因可能与松材线虫的发育速度、繁殖量、寿命、脂肪积累等表型相关。 |
| 关 键 词： | 松材线虫 灰葡萄孢菌 RNAi ncr基因 |
Functional analysis of the ncr gene from pine wood nematode Bursaphelenchus xylophilus using RNA interference |
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| WANG Dian-dong,CHEN Guo-hua,PAN Li-mei.Functional analysis of the ncr gene from pine wood nematode Bursaphelenchus xylophilus using RNA interference[J].Acta Phytopathologica Sinica,2018,48(4):518-526. | |
| Authors: | WANG Dian-dong CHEN Guo-hua PAN Li-mei |
| Institution: | Yangtze Normal University,Chongqing 408100,China; Institute of vegetables and flowers, Chinese Academy of Agricultural Sciences,Beijing 100081,China |
| Abstract: | The pine wilt disease (PWD) caused by Bursaphelenchus xylophilus is one of the most serious diseases that endanger a variety of pine trees. Since B. xylophilus is capable of rapid reproduction and spread, controlling PWD remains problematic worldwide, and effective approaches must be explored and established. RNAi technology is an important method to study the function of plant nematode genes. In this paper, the dsRNA expression vector for ncr gene (PDH-RH-NCR) was transformed into Botrytis cinerea by Agrobacturium-mediated transformation method. The phenotypes and ncr gene function were studied after fed on B. cinerea transformants for several generations. The results showed that the expression of ncr gene of B. xylophilus was significantly decreased 13 times after fed on B. cinerea transformants with PDH-RH-NCR expression vectors for 10 generations. The nematode development and reproduction were significantly reduced after the down-regulation of ncr gene. The percentage of adults developed from larvae individuals after 4 days was 61.75%, which was significantly lower than that of PDH-RH-sGFP (89.75%) and empty vector (88.75%) treatments. While the reproduction number after 9 days was 2 314 and 2 347 less than PDH-RH-sGFP and empty vector treatments, respectively. The average life span (20.9 d) and body fat mass (1.340 nmol) of 4th larvae after the down-regulation of ncr gene in PDH-RH-NCR treatment were significantly higher than PDH-RH-sGFP (15.5 d, 0.336 nmol) and empty vector treatments (15.2 d, 0.323 nmol). Therefore, the results showed that ncr gene could be related to the growth rate, reproduction capacity, life span and fat accumulation of B. xylophilus |
| Keywords: | Bursaphelenchus xylophilus Botrytis cinerea RNA interference ncr gene |
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