柑橘体细胞胚发生基因CsHB1特异肽段多克隆抗体的制备及其蛋白动态检测
发布时间：2018-12-15 17:04
【摘要】：【目的】构建柑橘HD-ZIP II转录因子CsHB1基因的原核表达系统,制备多克隆抗体,并检测抗体在‘伏令夏橙’胚性愈伤体胚诱导阶段中的特异性,为研究CsHB1基因在柑橘体细胞胚发生过程中的功能奠定基础。【方法】构建柑橘HD-ZIP II转录因子CsHB1基因的原核表达载体p GEX4T-CsHB1-N,转化大肠杆菌诱导目的融合蛋白的表达,并制备获得多克隆抗体anti-CsHB1-N。通过Western blot检测抗体在原核表达系统和柑橘体细胞胚诱导阶段的特异性,并分析体细胞胚诱导阶段CsHB1蛋白水平表达的动态变化。【结果】重组原核表达载体p GEX4T-CsHB1-N在Escherichia coli中高效表达出了分子质量约为49 ku的GST-CsHB1-N融合蛋白,并纯化获得多克隆抗体anti-CsHB1-N;经过Western blot分析表明,多克隆抗体可与‘伏令夏橙’愈伤体胚诱导阶段表达的目的蛋白特异结合;蛋白表达结果分析表明,在胚性愈伤组织中CsHB1蛋白呈现高的表达量,随着诱导培养的进行,呈现了先下降后上升的波动变化。【结论】多克隆抗体特异性好,可与‘伏令夏橙’胚性愈伤组织中目的蛋白特异性结合,可用于CsHB1基因功能分析。
[Abstract]:[objective] to construct the prokaryotic expression system of citrus HD-ZIP II transcription factor CsHB1 gene, to prepare polyclonal antibody, and to detect the specificity of the antibody in embryogenic callus induction stage. In order to study the function of CsHB1 gene in citrus somatic embryogenesis. [methods] the prokaryotic expression vector p GEX4T-CsHB1-N, of citrus HD-ZIP II transcription factor CsHB1 gene was constructed to express the fusion protein induced by Escherichia coli. The polyclonal antibody anti-CsHB1-N. was prepared. The specificity of antibody in prokaryotic expression system and citrus somatic embryogenesis was detected by Western blot. The dynamic changes of CsHB1 protein expression during somatic embryogenesis were analyzed. [results] the recombinant prokaryotic expression vector p GEX4T-CsHB1-N efficiently expressed GST-CsHB1-N fusion protein with molecular weight of about 49 ku in Escherichia coli. The polyclonal antibody anti-CsHB1-N; was purified and purified. Western blot analysis showed that the polyclonal antibody could specifically bind to the target protein expressed in the callus embryogenic stage of 'Fulingxia orange'. The results of protein expression showed that the expression of CsHB1 protein was high in embryogenic callus. With the development of induction and culture, the expression of CsHB1 protein decreased first and then increased. [conclusion] the polyclonal antibody has good specificity. It can specifically bind to the target protein in embryogenic callus of 'Volinga orange' and can be used for functional analysis of CsHB1 gene.
【作者单位】： 武汉生物工程学院应用生物技术研究中心;华中农业大学园艺林学学院;
【基金】：国家自然科学基金青年基金(31201611) 湖北省自然科学基金(2016CFB390)
【分类号】：S666
本文编号：2381002
[Abstract]:[objective] to construct the prokaryotic expression system of citrus HD-ZIP II transcription factor CsHB1 gene, to prepare polyclonal antibody, and to detect the specificity of the antibody in embryogenic callus induction stage. In order to study the function of CsHB1 gene in citrus somatic embryogenesis. [methods] the prokaryotic expression vector p GEX4T-CsHB1-N, of citrus HD-ZIP II transcription factor CsHB1 gene was constructed to express the fusion protein induced by Escherichia coli. The polyclonal antibody anti-CsHB1-N. was prepared. The specificity of antibody in prokaryotic expression system and citrus somatic embryogenesis was detected by Western blot. The dynamic changes of CsHB1 protein expression during somatic embryogenesis were analyzed. [results] the recombinant prokaryotic expression vector p GEX4T-CsHB1-N efficiently expressed GST-CsHB1-N fusion protein with molecular weight of about 49 ku in Escherichia coli. The polyclonal antibody anti-CsHB1-N; was purified and purified. Western blot analysis showed that the polyclonal antibody could specifically bind to the target protein expressed in the callus embryogenic stage of 'Fulingxia orange'. The results of protein expression showed that the expression of CsHB1 protein was high in embryogenic callus. With the development of induction and culture, the expression of CsHB1 protein decreased first and then increased. [conclusion] the polyclonal antibody has good specificity. It can specifically bind to the target protein in embryogenic callus of 'Volinga orange' and can be used for functional analysis of CsHB1 gene.
【作者单位】： 武汉生物工程学院应用生物技术研究中心;华中农业大学园艺林学学院;
【基金】：国家自然科学基金青年基金(31201611) 湖北省自然科学基金(2016CFB390)
【分类号】：S666
【相似文献】
相关期刊论文 前3条
1 李桂芬;马洁;陈红运;李明福;陈笑瑜;丁建云;;黄瓜绿斑驳花叶病毒多克隆抗体制备及检测应用[J];河南农业科学;2007年11期
2 张红星;朱本忠;张文;谢远宏;李欣;罗云波;;番茄乙烯信号转录因子LeERF1多克隆抗体制备及应用[J];中国食品学报;2008年01期
3 张红星;芮玉奎;张文;谢远宏;杨祥龙;尹胜;朱本忠;罗云波;;番茄乙烯信号转录因子LeERF2多克隆抗体的制备[J];生物技术通讯;2006年06期
，本文编号：2381002
本文链接：https://www.wllwen.com/kejilunwen/jiyingongcheng/2381002.html
最近更新
教材专著
