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1 518bp,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pFGC-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。
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 ofKCS 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.
出处
《作物研究》
2012年第3期213-218,共6页
Crop Research
基金
国家973项目(2006CB101603)
