抗吡虫啉苹果绵蚜nAChR亚基基因Elα2和Elα5的序列分析
毕业论文关键词：苹果绵蚜；吡虫啉；抗药性；烟碱型乙酰胆碱受体；氨基酸多态性
Analysis on sequences of nAChR subunit genes Elα2 and Elα5 from the imidacloprid-resistant populations of Eriosoma lanigerum
Abstract：Eriosoma lanigerum (Hausmann) is an important quarantine insect on fruit trees, which damaged seriously in the recent years. The previous results indicated that some field populations of E. lanigerum from China had evolved extremely high level of resistance to imidacloprid due to its unreasonable use. Imidacloprid is the first developed neonicotinoid insecticide, which acts on nicotinic acetylcholine receptors of insects. At present, resistance studies related to neonicotinoid insecticides are mainly focused on both metabolic resistance and target resistance. To elucidate the resistance mechanism of E. lanigerum to imidacloprid, the cloning and analysis of sequences of two nicotinic acetylcholine receptor subunits genes, Elα2 and Elα5, from E. lanigerum were carried out in the current study. The results showed that some amino acid polymorphisms were detected specifically in the sequence of Elα2 and Elα5 from different resistant populations, which is possibly associated with the resistance of E. lanigerum to imidacloprid.
Key words：Eriosoma lanigerum; imidacloprid；resistance；nAChR; amino acid polymorphism
目 录
摘要1
关键词1
Abstract1
Key words1
引言1
1 材料与方法2
1.1供试蚜虫 2
1.2 主要试剂和仪器2
1.2.1 主要试剂2
1.2.2 主要仪器2
1.3 总RNA提取2
1.3.1 总RNA提取2
1.3.2 苹果绵蚜总RNA质量检测3
1.4引物3
1.5 PCR扩增3
1.6 PCR产物回收与纯化3
1.7 质粒载体的连接与转化4
1.7.1 连接反应4
1.7.2 转化反应与单克隆培养4
1.7.3 挑斑4
1.8 序列分析4
2 苹果绵蚜烟碱型乙酰胆碱受体基因的点突变分析5
2.1不同苹果绵蚜种群Elα2和Elα5的克隆5
2.2 不同地区苹果绵蚜的Elα2和Elα5氨基酸多态性分析6
2.3 讨论与分析9
致谢10
参考文献10
抗吡虫啉苹果绵蚜nAChR亚基基因Elα2和Elα5的序列分析
引言
苹果绵蚜Eriosoma lanigerum（Hausmann）是一种重要的检疫性有害昆虫，在我国分布广泛，对苹果树危害严重【1】。主要寄生于苹果树枝干的新梢、伤口、叶腋、果梗、萼洼以及地下根部和根基等处为害，通过吸取树汁，从而削弱树势继而影响苹果树的生长、发育和花芽分化、影响苹果产量和质量【2】。
吡虫啉（Imidacloprid）是第一个开发出来的新烟碱类杀虫剂（Neonicotinonids），新烟碱类杀虫剂的作用靶标为昆虫烟碱型乙酰胆碱受体（nAChRs）【3】。由于长期频繁的使用，据不完全统计，已有包括烟粉虱、稻飞虱、烟蚜【4】等多种农业害虫对吡虫啉产生了抗性，许多研究表明，nAChR亚基氨基酸残基的缺失或者替换会导致昆虫抗性水平的改变【5-7】。Liu等（2005）在褐飞虱nAChRα1（Nlα1）亚基、nAChRα3（Nlα3）亚基中发现与吡虫啉抗性相关的突变位点，结果表明该突变可显著影响吡虫啉对褐飞虱Nlα1亚基的结合能力【8】。Bass等（2011）发现，桃蚜的nAChRβ1亚基的Loop环上存在突变使得桃蚜对吡虫啉的抗性水平升高【9】。Koo等（2014）也发现棉蚜nAChRβ1亚基的突变是导致抗性产生的重要原因【10】。
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