| 杜仲糖基转移酶基因EuCGT1克隆及序列分析 |
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| 引用本文： | 李永捷,;董旋,;赵德刚.杜仲糖基转移酶基因EuCGT1克隆及序列分析[J].贵州大学学报(农业与生物科学版),2014(4):27-35. |
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| 作者姓名： | 李永捷 ;董旋 ;赵德刚 |
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| 作者单位： | [1]贵州大学生命科学学院,山地植物资源保护与种质创新省部共建教育部重点实验室,贵州贵阳550025; [2]贵州大学农业生物工程研究院,贵州省农业生物工程重点实验室,贵州贵阳550025; [3]贵州大学绿色农药与农业生物工程国家重点实验室培养基地,贵州贵阳550025 |
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| 基金项目： | 国家863计划“特色植物功能基因组学研究与应用”子项目“特色林木功能基因组学研究与应用”子课题“杜仲功能基因组研究与应用”（2013AA102605-05）; 国家转基因生物新品种培育重大科技专项“安全转基因技术研究”子项目“保障转基因生物安全的外源基因清除技术研究”（2014ZX08010088-0022） |
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| 摘 要： | 本研究以杜仲(Eucommia ulmoides)7月雌株新生幼枝的树皮为材料,根据杜仲转录组测序数据中筛选的一个糖基转移酶基因(CGT)片段信息设计引物,采用SMART RACE技术,克隆了5’-端1432bp和3’-端535 bp cDNA片段,测序拼接后获得全长1 583 bp的杜仲CGT的cDNA序列。分析结果表明,该cDNA开放阅读框长1 464 bp,编码487个氨基酸,命名为EuGT基因。EuGT在NCBI中经blastn比对,显示序列与甜橙和草莓的UDP-葡萄糖基转移酶基因同源性分别为72%和67%;推测的氨基酸序列经blastp和CDD比对,显示序列与UDP-葡萄糖类黄酮3-O-糖基转移酶-6同源,其中含有UDP-葡萄糖基转移酶的保守域(pfam00201),初步推测EuCGT1蛋白属于糖基转移酶家族。本研究克隆的EuGT基因是首次从杜仲中克隆的糖基转移酶家族基因。以EuCGT1基因构建的植物表达载体pSH-35S-EuCGT1采用农杆菌介导法遗传转化烟草,获得了含EuCGT1基因的转基因烟株,移栽生长1个月的转基因烟草植株在表型上与野生型烟草无明显差异。
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| 关 键 词： | 杜仲 糖基转移酶 RACE技术 |
Cloning and Sequence Analysis of Glucosyltransferase Gene EuCGT1 in Eucommia ulmoides Olive |
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| Institution: | LI Yong-jie, DONG Xuan, ZHAO De-gang ( 1. The Key Laboratory of Plant Resources Conservation and C, ermplasm Innovation in Mountainous Region, Ministry of Education, College of Life Sciences, Guizhou University, Guiyang Guizhou 550025, China ; 2. Guizhou Key Lab of Agro - Bioengineering, Institute of Agro - Bioengineering, Guizhou University, Guiyang Guizhou 55(25 , China; 3. State Key Laboratory Breeding Base of Green Pesticide and Agricultural Bioengineering, Guizhou University, Guiyang Guizhou 550025, China) |
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| Abstract: | The cDNA sequence of glucosyltransferase gene （EuCGT1） was cloned from the total RNA of Eucommia ulmoides bark using the method of SMART-RACE. The results revealed that the EuCGT1 was 1 583 bp in length, containing an open reading frame （ ORF） of 1 464 bp encoding a protein of 487 amino acids. EuCGT1 sequence shared 72% and 67% DNA homology with the UDP-GT gene sequence of Citrus sinensis and Fragaria x ananas-sa, respectively. The conserved domains database （ CDD） search suggested the EuCGT1 had similar domain to pfam00201, as the UDP- Glucosyltransferase proteins. Dicot expressionas vector pSH-35S-EuCGT1 was con-structed and transformed into Nicotiana tabacum L. via mediated with Agrobacterium tumefaciens. Seven transgenic tobacco lines were obtained after GUS histochemical staining and PCR molecular detection. The transgenic tobacco demonstrated similar phenotypes as the wild types after one month growth. |
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| Keywords: | Eucommia ulmoides Olive Glucosyltransferase SMART-RACE |
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