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斜纹夜蛾触角气味结合蛋白基因SlitGOBP2的克隆与序列分析 被引量:5
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作者 钟国华 李苗孟 +2 位作者 胡美英 罗倩 胡黎明 《华中农业大学学报》 CAS CSCD 北大核心 2008年第5期578-584,共7页
应用RT-PCR扩增结合RACE技术克隆获得斜纹夜蛾触角普通气味结合蛋白2(GOBP2)基因Slit-GOBP2全序列。SlitGOBP2的阅读框全长489bp,编码162个氨基酸残基,预测分子质量和等电点分别为18.16ku和5.43。cDNA和氨基酸序列GenBank登录号分别为EF... 应用RT-PCR扩增结合RACE技术克隆获得斜纹夜蛾触角普通气味结合蛋白2(GOBP2)基因Slit-GOBP2全序列。SlitGOBP2的阅读框全长489bp,编码162个氨基酸残基,预测分子质量和等电点分别为18.16ku和5.43。cDNA和氨基酸序列GenBank登录号分别为EF159979和ABM54824。SlitGOBP2氨基酸序列含6个保守的半胱氨酸位点,具有GOBP2的典型特征,与16种鳞翅目昆虫GOBP2同源性为94%~75%;编码蛋白呈酸性,整个分子呈亲水性,在第40~60位和100位以后的氨基酸表现亲脂性,蛋白质二级结构主要由α螺旋构成。氨基酸序列系统进化树分析表明,SlitGOBP2与草地夜蛾Spodoptera frugiperda和甘蓝夜蛾Mamestra brassicae的GOBP2同属于一个分支,一致性分别为94.4%和92.0%。昆虫GOBP2分子在体现保守性的同时也体现了进化性。 展开更多
关键词 斜纹夜蛾 普通气味结合蛋白2 基因克隆 序列分析
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Cloning and Sequencing of a Gene Encoding GOBP2 in the Antenna of Spodoptera exigua
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作者 WANG Gui-rong, GUO Yu-yuan, XU Guang and WU Kong-ming( The Institute of Plant Protection , Chinese Academy of Agricultural Sciences , Beijing 100094 ) 《Agricultural Sciences in China》 CAS CSCD 2002年第2期200-206,共7页
A pair of degenerate primers was designed, based on the comparison of five insects' GOBP2 gene sequences reported previously. A specific band (about 400bp in length) was amplified from cDNA of Spodoptera exigua an... A pair of degenerate primers was designed, based on the comparison of five insects' GOBP2 gene sequences reported previously. A specific band (about 400bp in length) was amplified from cDNA of Spodoptera exigua antenna and another specific band (about 2kb in length) was amplified from genomic DNA. The two segments were cloned into T-easy vector, respectively. Results of sequencing and structural analysis showed that the full-length of GOBP2Sexi ORF is 426bp, 141 amino acid residues were encoded. The predicted MW and pI are 16.07ku and 5.09 .respectively. There are six conservative Cys locus in the sequence, which is the typical characteristic of OBPs. GOBP2Sexi gene was inserted by two introns between amino acid residue 22 and 23 and between 82 and 83. The length of two introns is 160bp and 1403bp. Results of Northern blot showed that GOBP2 gene expressed specifically in the antenna of Spodoptera exigua, and the expression level is nearly equal in the antenna of male and female moths. The sequence was deposited in GenBank/EMBL and the accession number is AJ294809. 展开更多
关键词 Spodoptera exigua ANTENNA gobp2 Gene cloning
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小菜蛾普通气味结合蛋白2基因的原核表达与多克隆抗体制备 被引量:2
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作者 徐红星 陈东旭 +1 位作者 张雅林 王敦 《环境昆虫学报》 CSCD 北大核心 2010年第2期194-198,共5页
本实验提取羽化3d的小菜蛾Plutella xylostella成虫触角总RNA,反转录合成cDNA,以此为模板PCR扩增出小菜蛾普通气味结合蛋白2基因,大小为492bp,Blast结果显示与多种昆虫的GOBP2具有较高的同源性。将该基因克隆到表达载体pMAL-c4E中,转化... 本实验提取羽化3d的小菜蛾Plutella xylostella成虫触角总RNA,反转录合成cDNA,以此为模板PCR扩增出小菜蛾普通气味结合蛋白2基因,大小为492bp,Blast结果显示与多种昆虫的GOBP2具有较高的同源性。将该基因克隆到表达载体pMAL-c4E中,转化宿主菌TB1(DE3),获得单克隆重组质粒pMAL-c4E-GOBP2。IPTG成功诱导pMAL-c4E-GOBP2表达出约60kDa的融合蛋白。优化诱导条件为3mmol/L终浓度IPTG、6h,可获得大量可溶性蛋白。表达的融合蛋白通过亲和色谱法纯化、免疫新西兰大白兔制备多克隆抗体。ELISA分析表明制备的抗体效价达1:1.28×105。 展开更多
关键词 小菜蛾 gobp2 原核表达 多克隆抗体
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