[1]黄磊,彭英传,韩召军.大螟乙酰胆碱酯酶基因的克隆及其多态性分析[J].南京农业大学学报,2019,42(6):1050-1058.[doi:10.7685/jnau.201903046]
 HUANG Lei,PENG Yingchuan,HAN Zhaojun.Cloning and polymorphism analysis of acetylcholinesterase genes from Sesamia inferens[J].Journal of Nanjing Agricultural University,2019,42(6):1050-1058.[doi:10.7685/jnau.201903046]
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大螟乙酰胆碱酯酶基因的克隆及其多态性分析()
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《南京农业大学学报》[ISSN:1000-2030/CN:32-1148/S]

卷:
42卷
期数:
2019年6期
页码:
1050-1058
栏目:
植物科学
出版日期:
2019-11-15

文章信息/Info

Title:
Cloning and polymorphism analysis of acetylcholinesterase genes from Sesamia inferens
作者:
黄磊1 彭英传2 韩召军1
1. 南京农业大学植物保护学院/农作物生物灾害综合治理教育部重点实验室, 江苏 南京 210095;
2. 江西农业大学昆虫研究所, 江西 南昌 330045
Author(s):
HUANG Lei1 PENG Yingchuan2 HAN Zhaojun1
1. College of Plant Protection/Key Laboratory of Integrated Management of Crop Diseases and Pests, Ministry of Education, Nanjing Agricultural University, Nanjing 210095, China;
2. Institute of Entomology, Jiangxi Agricultural University, Nanchang 330045, China
关键词:
大螟乙酰胆碱酯酶基因克隆多态性
Keywords:
Sesamia inferensacetylcholinesterasegene cloningpolymorphism
分类号:
S435.112+.1
DOI:
10.7685/jnau.201903046
摘要:
[目的] 本研究旨在通过克隆大螟的乙酰胆碱酯酶基因ace,进行多态性分析,以便寻找抗药性相关的突变位点,为建立高通量药剂筛选方法和适宜的抗药性分子检测方法提供新途径。[方法] 利用PCR和cDNA末端快速扩增(RACE)技术克隆大螟的ace全长序列,通过分子特征和构建系统发育树对其进行验证。通过序列比对,分析不同地理种群大螟ace基因的多态性,寻找经典抗药性突变位点,同时测定相应种群的三唑磷抗性,联合分析相关的抗药性突变位点。[结果] 克隆获得了大螟2个ace基因的全长cDNA,分别将其命名为Sinace1Sinace2(GenBank登录号分别为MK641586和MK641585)。它们均含有乙酰胆碱酯酶家族的保守结构特征,系统发育树分析表明它们和鳞翅目近缘种的ace基因亲缘关系最近。关键靶标基因Sinace1在4个地理种群的97条基因序列中共发现了比率不同的107种核苷酸多态性位点和21种氨基酸突变类型,但未发现经典抗药性位点的突变。生物测定结果表明:不同地理种群的大螟对三唑磷的敏感性相差较小(1~5倍),对比分析也没有发现种群之间突变比率与抗性水平一致的突变位点。[结论] 大螟ace1基因尚未产生可测定的抗药性突变,但本研究率先克隆报道了大螟的2个ace基因全长序列,并揭示了大螟不同地理种群ace1基因的主要多态性位点。
Abstract:
[Objectives] This study tries to seek a new molecular approach substitute by cloning the acetylcholinesterase genes(ace)from the Sesamia inferens and analyzing their polymorphism and resistance related mutations. It provides a new way to establish high-throughput insecticide screening and suitable insecticide resistance molecular detection methods.[Methods] Acetylcholinesterase genes were cloned by polymerase chain reaction(PCR)and rapid amplification of cDNA ends(RACE),and confirmed by their molecular characteristics and phylogenetic tree analysis. The polymorphism frequencies were analyzed for the key gene ace1 among different geographical populations. The corresponding resistance levels to triazophos were also tested,and finally the resistance mutations were pursued.[Results] Two full-length cDNA sequences from S.inferens were cloned,characterized,and designated as Sinace1 and Sinace2(GenBank accession No.:MK641586 and MK641585),respectively. They both shared all the conserved structural features of the ace family,and the phylogenetic tree analysis indicated that they had the closest relationship with the corresponding ace genes from lepidopeterians. Polymorphism analysis of the 97 Sinace1 sequences showed 107 nucleotide polymorphism sites and 21 amino acid mutations all with varied frequencies among the four geographical populations. No resistant mutations previously reported in different ace genes were found. Four of the geographical populations were tested and showed that their sensitivities to triazophos varied within 5 times. Comparison analysis found no amino acid mutations with their occurrences consistent with triazophos resistance among the geographical populations.[Conclusions] No detectable resistance mutations have happened in Sinace1 of S.inferens. However,the two ace genes and the polymorphism firstly reported in this work should lay a foundation for the study of the resistance mechanism for organophosphate resistance,and technique development for selecting efficient insecticides by enzyme activity inhibition.

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备注/Memo

备注/Memo:
收稿日期:2019-03-21。
基金项目:国家科技基础资源调查专项(2018FY101100)
作者简介:黄磊,硕士研究生。
通信作者:韩召军,教授,主要从事昆虫生理生化与分子毒理学研究,E-mail:zjhan@njau.edu.cn。
更新日期/Last Update: 1900-01-01