| 柚CCoAOMT基因的克隆与超表达载体构建 |
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| 引用本文： | 徐玉,潘腾飞,潘东明.柚CCoAOMT基因的克隆与超表达载体构建[J].中国农学通报,2014,30(25):148-153. |
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| 作者姓名： | 徐玉 潘腾飞 潘东明 |
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| 作者单位： | 1. 福建农林大学2. 3. 福建农林大学园艺学院 |
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| 基金项目： | 国家科技支撑计划项目 高淀粉酿造高粱新品种选育与产业化示范;福建省种业创新与产业化工程项目、福建农林大学创新团队项目 |
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| 摘 要： | 为探明咖啡酰辅酶A-O-甲基转移酶（CCoAOMT）在柚Citrus maxima (Burm.) Merr.]果实发育中对木质素单体合成的影响，克隆了柚CCoAOMT基因ORF序列并构建超表达载体转化至农杆菌。采用Trizol法提取柚汁胞总RNA，经反转录得到cDNA，根据已知柚CCoAOMT片段序列，设计其ORF引物。PCR扩增得到柚CCoAOMT基因ORF序列，长度为744 bp，编码247个氨基酸，与龙眼(Dimocarpus longan)、麻疯树(Jatropha curcas)、杨树(Populus balsamifera)的CCoAOMT基因同源性大于90%。经软件预测，该酶定位于质膜的可能性最大，不具有跨膜结构。将柚CCoAOMT基因正向导入植物过表达载体p1301，位置在CaMV35S启动子的之后，并将载体转化至农杆菌EHA105。本研究得到的基因片段包含柚CCoAOMT基因ORF，成功构建植物超表达载体p1301-cmCCoAOMT，并导入农杆菌。含p1301-cm CCoAOMT的农杆菌株可转化多种植物受体，为深入研究该基因功能奠定了基础。
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| 关 键 词： | 抑制机制 抑制机制 |
| 收稿时间： | 2014/2/21 0:00:00 |
| 修稿时间： | 2014/5/24 0:00:00 |
Cloning of Caffeoyl-coenzyme A O-methyltransferase Gene from Citrus maxima (Burm.) Merr. and Its Plant Expressing Vector Construction |
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| Institution: | Xu Yu, Pan Tengfei, Pan Dongming (1College of Horticulture, Fujian Agricultural and Forestry University, Fuzhou 350002; 2.Institute of Postharvest Science and Technology of Horticultural Products, FAFU, Fuzhou 350002) |
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| Abstract: | To probe the effects of the Caffeoyl-coenzyme A O-methyltransferase on lignin monomer biosynthesis of pummelo Citrus maxima (Burm.) Merr.]. A cmCCoAOMT gene was cloned from pummelo juice sac. Total RNA was obtained by Trizol method, and was reverse transcribed as cDNA. A cmCCoAOMT1 gene was cloned by PCR with a primer pair designed from a CCoAOMT gene sequence. The ORF contained 744 base pairs coding 247 amino acids. The sequence homology with Populus balsamifera, Jatropha curcas and Dimocarpus longan exceeded 90% respectively. Prediction result indicated that cmCCoAOMT1 was located on ER with possibility 0.595. A mediate vector p1301-AO was employed to add a digestion site Apa I into p1301, and then p1301- CCoAOMT1 with Apa I was built. |
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| Keywords: | expression vector construction |
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