淡色库蚊ATP合酶B亚基基因克隆和序列分析及其与溴氰菊酯抗性的关系

doi:10.16380/j.kcxb.2017.08.006

›› 2017, Vol. 60 ›› Issue (8): 900-905.doi: 10.16380/j.kcxb.2017.08.006

淡色库蚊ATP合酶B亚基基因克隆和序列分析及其与溴氰菊酯抗性的关系

王卫杰^{1, 2,*}, 刘新颖³, 方福瑾^{2, 4}, 过琴², 王贺¹, 齐莉莉¹, 赵文爱¹

( 1. 河北医科大学病原生物学教研室, 石家庄 050017; 2. 南京医科大学病原生物学系, 南京 211166; 3. 保定市第二中心医院血液肾病科, 河北涿州 072750; 4. 蚌埠市第三人民医院检验科, 安徽蚌埠 233300 )

出版日期:2017-08-20 发布日期:2017-08-20

Cloning and sequence analysis of ATP synthase B subunit gene and its association with deltamethrin resistance in Culex pipiens pallens (Diptera: Culicidae)

WANG Wei-Jie^1,2,*,LIU Xin-Ying³, FANG Fu-Jin^2,4, GUO Qin², WANG He¹, QI Li-Li¹, ZHAO Wen-Ai¹

(1. Department of Pathogen Biology, Hebei Medical University, Shijiazhuang 050017, China; 2. Department of Pathogen Biology, Nanjing Medical University, Nanjing 211166, China; 3. Department of Hematology and Nephrology, Second Central Hospital of Baoding City, Zhuozhou, Hebei 072750, China; 4. Department of Clinical Laboratory, the Third People’s Hospital of Bengbu, Bengbu, Anhui 233000, China)

Online:2017-08-20 Published:2017-08-20

摘要/Abstract

摘要： 【目的】克隆淡色库蚊Culex pipiens pallens ATP合酶B亚基基因编码区序列，并进行生物信息学分析，研究其与溴氰菊酯抗性关系。【方法】通过PCR方法扩增ATP合酶B亚基基因编码区序列；利用生物信息学网站在线分析工具，预测ATP合酶B亚基基因编码蛋白的理化性质和功能特征；通过实时定量PCR方法比较ATP合酶B亚基基因在室内筛选的淡色库蚊溴氰菊酯敏感品系和抗性品系中的表达，并在现场种群溴氰菊酯敏感和抗性个体中做进一步验证。【结果】成功克隆淡色库蚊ATP合酶B亚基基因编码区序列(GenBank登录号: KY783434)，长717 bp，编码238个氨基酸。生物信息学分析表明，其编码蛋白理论相对分子量为26.96 kD，等电点为8.97，在第70-231位氨基酸具有ATP合酶B亚基结构域，未发现信号肽和跨膜区。实时定量PCR结果显示，ATP合酶B亚基基因在室内筛选的淡色库蚊抗溴氰菊酯品系和现场种群溴氰菊酯抗性个体中表达量均上调。【结论】本研究获得了淡色库蚊ATP合酶B亚基基因编码区序列，进行了生物信息学分析，并证实其在溴氰菊酯抗性个体中高表达，为进一步研究该基因在蚊抗药性中的作用奠定了基础。

关键词: 淡色库蚊, ATP合酶B亚基, 抗药性, 溴氰菊酯, 生物信息学

Abstract: 【Aim】 This study aims to clone the coding sequence of ATP synthase B subunit gene, to perform bioinformatics analysis for the coding sequence and to characterize its association with deltamethrin resistance in Culex pipiens pallens. 【Methods】 The coding sequence of ATP synthase B subunit gene in C. pipiens pallens was amplified by PCR method. The physical-chemical properties and functional characteristics of the encoded protein were predicted by employing the web-based bioinformatics analysis. The expression of ATP synthase B subunit gene was compared between the deltamethrin-susceptible and-resistant strains selected in the laboratory by quantitative real-time PCR method, and between the deltamethrin-susceptible and -resistant individuals of the field populations for further verification. 【Results】 The coding sequence of ATP synthase B subunit gene (GenBank accession no.: KY783434) was cloned, which contains 717 nucleotides encoding 238 amino acids. Bioinformatics analysis showed that its encoded protein has a theoretical relative molecular weight of 26.96 kD and the isoelectric point value of 8.97 approximately. It was predicted that ATP synthase B subunit contains a structural domain (amino acids 70-231) of ATP-synthase_B, but lacks signal peptide and transmembrane domain. Quantitative real-time PCR results revealed that the expression level of ATP synthase B subunit gene was up-regulated in the deltamethrin-resistant strain as compared with that in the deltamethrin-susceptible strain selected in the laboratory, and also in the deltamethrin-resistant individuals of the field populations as compared with that in the deltamethrin-susceptible individuals of the field populations. 【Conclusion】 This study obtained the coding sequence of ATP synthase B subunit gene, performed bioinformatics analysis for the sequence, and confirmed its overexpression in the deltamethrin-resistant individuals, providing the basis for further studying its roles in insecticide resistance.

Key words: Culex pipiens pallens, ATP synthase B subunit, insecticide resistance, deltamethrin, bioinformatics

王卫杰, 刘新颖, 方福瑾, 过琴, 王贺, 齐莉莉, 赵文爱. 淡色库蚊ATP合酶B亚基基因克隆和序列分析及其与溴氰菊酯抗性的关系[J]. , 2017, 60(8): 900-905.

WANG Wei-Jie, LIU Xin-Ying, FANG Fu-Jin, GUO Qin, WANG He, QI Li-Li, ZHAO Wen-Ai. Cloning and sequence analysis of ATP synthase B subunit gene and its association with deltamethrin resistance in Culex pipiens pallens (Diptera: Culicidae)[J]. , 2017, 60(8): 900-905.

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链接本文: http://www.insect.org.cn/CN/10.16380/j.kcxb.2017.08.006

http://www.insect.org.cn/CN/Y2017/V60/I8/900

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淡色库蚊ATP合酶B亚基基因克隆和序列分析及其与溴氰菊酯抗性的关系

Cloning and sequence analysis of ATP synthase B subunit gene and its association with deltamethrin resistance in Culex pipiens pallens (Diptera: Culicidae)

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