| HARDY基因的克隆、原核表达及其植物表达载体的构建 |
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| 引用本文： | 马玲珑,林国梁,赖 勇,马小乐,孟亚雄,李葆春,王化俊.HARDY基因的克隆、原核表达及其植物表达载体的构建[J].麦类作物学报,2014,34(3):292-297. |
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| 作者姓名： | 马玲珑 林国梁 赖 勇 马小乐 孟亚雄 李葆春 王化俊 |
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| 作者单位： | （1．甘肃省作物遗传改良与种质创新重点实验室/甘肃省干旱生境作物学重点实验室，甘肃兰州 730070；2．甘肃农业大学农学院，甘肃兰州 730070； 3．甘肃农业大学生命科学技术学院，甘肃兰州 730070） |
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| 基金项目： | 甘肃省农业科技创新项目；甘肃省科技厅重大专项（0801NKDA013）；甘肃省农业科技成果转化资金计划项目（1006NCNA120）。 |
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| 摘 要： | 为了给转HARDY(HRD)基因小麦研究奠定基础,利用PCR技术从拟南芥中克隆HRD基因,进行生物信息学分析,预测其编码的蛋白质结构与功能,构建原核表达载体pET28a(+)-HRD,转化大肠杆菌BL21(DE3),IPTG诱导蛋白的表达,同时构建pBin-HRD植物表达载体。测序分析结果表明,克隆的HRD与NCBI发布的拟南芥核苷酸序列的同源性为99%,其开放阅读框长555bp,可编码184个氨基酸,具有许多重要功能位点;成功构建了原核表达载体pET28a(+)-HRD和植物表达载体pBin-HRD,诱导表达出大小约为23.3kD的蛋白,与理论值相近。
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| 关 键 词： | HARDY（HRD） 基因克隆 原核表达 植物表达载体 |
Cloning, Prokaryotic Expression and Construction of Plant Expression Vector for HARDY Gene |
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| MA Linglong,LIN Guoliang,LAI Yong,MA Xiaole,MENG Yaxiong,LI Baochun,WANG Huajun.Cloning, Prokaryotic Expression and Construction of Plant Expression Vector for HARDY Gene[J].Journal of Triticeae Crops,2014,34(3):292-297. |
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| Authors: | MA Linglong LIN Guoliang LAI Yong MA Xiaole MENG Yaxiong LI Baochun WANG Huajun |
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| Abstract: | To lay a foundation for studying HARDY(HRD) gene in transgenic wheat,HRD was cloned from Arabidopsis thaliana by PCR, and the structure and function of its encoded protein was predicted by bioinformatics analysis. Prokaryotic expression vector pET28a(+) HRD was constructed and transformed into E.coli BL21(DE3) to induce protein expression with IPTG, and plant expression vector pBin HRD was constructed. The sequence analysis showed that the nucleotide sequence of HRD shared 99% similarity with HRD of Arabidopsis thaliana published in NCBI, and its full length was 555 bp, which encoded 184 amino acids and containing many important functional sites. Prokaryotic expression vector pET28a(+) HRD and plant expression vector pBin HRD was constructed successfully, and a recombinant protein of approximately 23.3 kD was expressed in the BL21 (DE3), which was similar to the theoretical value. |
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| Keywords: | HARDY(HRD) Gene cloning Prokaryotic expression Plant expression vector |
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