柑橘大实蝇化学感受蛋白基因BminCSP3的克隆、表达及结合特征分析
S666%S436.661; [目的]探究在柑橘大实蝇成虫触角高表达的化学感受蛋白BminCSP3与气味配体的结合特性,以期为揭示柑橘大实蝇嗅觉识别的分子机制提供一定理论依据.[方法]通过RT-PCR克隆BminCSP3基因,并对其进行生物信息学分析;以原核表达系统和镍柱体外表达纯化得到BminCSP3蛋白;利用荧光竞争结合实验检测BminCSP3与14种气味配体的结合特性.[结果]BminCSP3开放阅读框全长为474 bp,编码157个氨基酸,含有4个半胱氨酸残基,为典型的化学感受蛋白;预测其等电点为5.41,信号肽由22个氨基酸组成,无跨膜结构域,亲水性总平均值为-0.397,是亲水性蛋...
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| Published in | 果树学报 Vol. 39; no. 3; pp. 343 - 351 |
|---|---|
| Main Authors | , , , , , , |
| Format | Journal Article |
| Language | Chinese |
| Published |
长江大学农学院·长江大学昆虫研究所,湖北荆州434025%长江大学生命科学学院,湖北荆州434025
10.03.2022
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| Subjects | |
| Online Access | Get full text |
| ISSN | 1009-9980 |
| DOI | 10.13925/j.cnki.gsxb.20210336 |
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| Abstract | S666%S436.661; [目的]探究在柑橘大实蝇成虫触角高表达的化学感受蛋白BminCSP3与气味配体的结合特性,以期为揭示柑橘大实蝇嗅觉识别的分子机制提供一定理论依据.[方法]通过RT-PCR克隆BminCSP3基因,并对其进行生物信息学分析;以原核表达系统和镍柱体外表达纯化得到BminCSP3蛋白;利用荧光竞争结合实验检测BminCSP3与14种气味配体的结合特性.[结果]BminCSP3开放阅读框全长为474 bp,编码157个氨基酸,含有4个半胱氨酸残基,为典型的化学感受蛋白;预测其等电点为5.41,信号肽由22个氨基酸组成,无跨膜结构域,亲水性总平均值为-0.397,是亲水性蛋白.荧光竞争结合实验结果表明,BminCSP3与供试14种气味配体的亲和力均较弱,其Ki值均大于20 pmol·L-1.一般认为Ki值大于20 μmol·L-1,说明蛋白与配体分子间的亲和力较弱.[结论]BminCSP3虽然在柑橘大实蝇成虫触角中显著上调表达,但可能不参与柑橘大实蝇成虫的嗅觉识别过程;或本试验所用配体并非BminCSP3最适配体,应扩大配体库进一步验证BminCSP3的嗅觉功能. |
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| AbstractList | S666%S436.661; [目的]探究在柑橘大实蝇成虫触角高表达的化学感受蛋白BminCSP3与气味配体的结合特性,以期为揭示柑橘大实蝇嗅觉识别的分子机制提供一定理论依据.[方法]通过RT-PCR克隆BminCSP3基因,并对其进行生物信息学分析;以原核表达系统和镍柱体外表达纯化得到BminCSP3蛋白;利用荧光竞争结合实验检测BminCSP3与14种气味配体的结合特性.[结果]BminCSP3开放阅读框全长为474 bp,编码157个氨基酸,含有4个半胱氨酸残基,为典型的化学感受蛋白;预测其等电点为5.41,信号肽由22个氨基酸组成,无跨膜结构域,亲水性总平均值为-0.397,是亲水性蛋白.荧光竞争结合实验结果表明,BminCSP3与供试14种气味配体的亲和力均较弱,其Ki值均大于20 pmol·L-1.一般认为Ki值大于20 μmol·L-1,说明蛋白与配体分子间的亲和力较弱.[结论]BminCSP3虽然在柑橘大实蝇成虫触角中显著上调表达,但可能不参与柑橘大实蝇成虫的嗅觉识别过程;或本试验所用配体并非BminCSP3最适配体,应扩大配体库进一步验证BminCSP3的嗅觉功能. |
| Author | 王兆祥 桂连友 杨岭 陈剑 张国辉 田晓丽 王福莲 |
| AuthorAffiliation | 长江大学农学院·长江大学昆虫研究所,湖北荆州434025%长江大学生命科学学院,湖北荆州434025 |
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| Author_FL | ZHANG Guohui CHEN Jian TIAN Xiaoli YANG Ling GUI Lianyou WANG Zhaoxiang WANG Fulian |
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| Keywords | 蛋白纯化 原核表达 化学感受蛋白 荧光竞争结合实验 柑橘大实蝇 |
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