| 碎米荠CarKCS基因全长CDS克隆、序列分析及表达载体构建 |
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| 引用本文： | 杨慧,魏解冰,阮颖,刘春林.碎米荠CarKCS基因全长CDS克隆、序列分析及表达载体构建[J].作物研究,2012,26(3):213-218. |
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| 作者姓名： | 杨慧 魏解冰 阮颖 刘春林 |
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| 作者单位： | 1. 作物种质资源创新与利用国家重点实验室培育基地,长沙410128;湖南农业大学农学院
2. 作物种质资源创新与利用国家重点实验室培育基地,长沙410128;湖南农业大学生物科学与技术学院,长沙410128 |
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| 基金项目： | 国家973项目(2006CB101603) |
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| 摘 要： | 根据KCS基因的保守性设计引物,以高神经酸含量的碎米荠叶片DNA为模板(KCS基因无内含子),克隆碎米荠超长链脂肪酸合成的限速酶β-酮脂酰- CoA合酶(KCS)基因全长CDS序列,命名为CarKCS.Blast 比对及序列分析表明,目的片段序列和GenBank上报道的拟南芥KCS18序列同源性达到87％.CarKCS全长1518bp,不含内含子,编码505个氨基酸.生物信息学分析表明,所有的酶功能活性位点的氨基酸都存在,在N端都有两个,C端有一个高度疏水的跨膜结构域；CarKCS与KCS18同属于KCS基因家族的FAE1 - like亚家族.将目的片段连接到pFCC -5941.nap表达载体,经PCR和酶切检测,证明已成功构建了pNapin- CarKCS载体.
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| 关 键 词： | 碎米荠 β-酮脂酰-CoA合酶(KCS)基因 序列分析 表达载体构建 |
Clone,Sequence Analysis and Expression Vector Construction of CarKCSGene Full-length CDS in Cardamine hirsute |
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| YANG Hui
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WEI Jie-bing
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RUAN Ying
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LIU Chun-lin.Clone,Sequence Analysis and Expression Vector Construction of CarKCSGene Full-length CDS in Cardamine hirsute[J].Crop Research,2012,26(3):213-218. |
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| Authors: | YANG Hui WEI Jie-bing RUAN Ying LIU Chun-lin |
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| Institution: | 1,2*(1 Pre-State Key Laboratory of Crop Germplasm Innovation and Utilization,Hunan Agricultural University,Changsha, Hunan 410128,China;2 College of Agronomy,Hunan Agricultural University,Changsha,Hunan 410128,China; 3 College of Bioscience and Biotechnology,Hunan Agricultural University,Changsha,Hunan 410128,China) |
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| Abstract: | In this study,according to the conservative sequence of β-ketoacyl-CoA synthase(KCS) genes,we designed the primers and cloned a full-length CDS of KCS gene from Cardamine hirsute,named CarKCS.Blast analysis showed that the homology between the cloned sequence and Arabidopsis thaliana KCS18 sequence in GenBank was up to 87%.The CarKCS contained 1518 bp and coded 505 amino acids.Bioinformatic analysis showed that CarKCS gene included all amino-acid residues of the enzyme active site and there were two in the N-terminal and one in the C-terminal highly hydrophobic transmembrane domains.CarKCS liked KCS18 belonged to FAE1-like subfamily of KCS gene family.PCR and restriction enzyme digestion demonstrated that we had successfully constructed pNapin-CarKCS vector,which provided good foundation for further study of CarKCS gene function. |
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| Keywords: | Cardamine hirsute β-ketoacyl-CoA synthase(KCS) gene Sequence analysis Construction of expression vector |
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