珍稀细茎石斛LEAFY同源基因的克隆及在石斛药材鉴定中的意义  

Sequence analysis of LEAFY homologous gene from Dendrobium moniliforme and application for identification of medicinal Dendrobium

作　　者：邢文瑞[1] 侯北伟[1] 管婧姣[1] 罗晶[1] 丁小余[1] 

机构地区：[1]南京师范大学生命科学学院,江苏南京210023

出　　处：《药学学报》2013年第4期597-603,共7页Acta Pharmaceutica Sinica

基　　金：国家自然科学基金资助项目(31170300);高等学校博士学科点专项科研基金资助项目(20093207110006)

摘　　要：本文基于兰科已有的LEAFY(LFY)同源基因保守区设计引物,对细茎石斛(Dendrobium moniliforme(L.)Sw.)的该基因进行克隆及序列分析。通过普通PCR及Tail-PCR技术得到LFY同源基因——DenLFY,全长共3 575 bp,包含3个外显子和2个内含子。将DenLFY基因外显子序列提交到NCBI进行BLAST比对,发现该基因与兰科其他植物同源基因序列相似度很高,均达80%以上,其中与蝴蝶兰(Phalaenopsis hybrid cultivar)LFY同源基因(PhalLFY)和文心兰(Oncidium Gower Ramsey)LFY同源基因相似度分别达84%和90%。应用MP的方法,根据不同物种LFY同源基因进行系统发生关系分析,发现细茎石斛与文心兰、蝴蝶兰亲缘关系最近。氨基酸序列比对分析结果表明,该基因C端编码的氨基酸相对更为保守。从DenLFY基因外显子和内含子单独分析来看,外显子的获得及其编码氨基酸的推测对于LFY基因功能的研究是一个开端,而内含子2在细茎石斛和铁皮石斛(Dendrobium officinale Kimura et Migo)中存在长度差异,在石斛药材鉴定方面有着很大的应用潜力。The LEAFY （LFY） homologous gene of Dendrobium moniliforme （L.） Sw. was cloned by new primers which were designed based on the conservative region of known sequences of orchid LEAFY gene. Partial LFY homologous gene was cloned by common PCR, then we got the complete LFY homologous gene DenLFY by Tail-PCR. The complete sequence of DenLFY gene was 3 575 bp which contained three exons and two introns. Using BLAST method, comparison analysis among the exon of LFYhomologous gene indicted that the DenLFY gene had high identity with orchids LFY homologous, including the related fragment of PhalLFY （84%） in Phalaenopsis hybrid cultivar, LFY homologous gene in Oncidium （90%） and in other orchid （over 80%）. Using MP analysis, Dendrobium is found to be the sister to Oncidium and Phalaenopsis. Homologous analysis demonstrated that the C-terminal amino acids were highly conserved. When the exons and introns were separately considered, exons and the sequence of amino acid were good markers for the function research of DenLFY gene. The second intron can be used in authentication research of Dendrobium based on the length polymorphism between Dendrobium moniliforme and Dendrobium officinale.

关 键 词：细茎石斛 LEAFY同源基因 Tail-PCR技术 序列分析 

分 类 号：R931[医药卫生—生药学]