萼脊兰SeAP1-like基因的克隆与表达载体构建  

Cloning and Construction of Expression Vector of SeAP1-like Gene from Sedirea japonica

作　　者：蒋素华[1] 袁秀云[1] 王洁琼[2] 武振江[2] 张国付[2] 崔波[1] 

机构地区：[1]郑州师范学院生物工程研究所,河南郑州450044 [2]河南农业大学生命科学学院,河南郑州450002

出　　处：《河南农业科学》2015年第1期105-109,共5页Journal of Henan Agricultural Sciences

基　　金：河南省重点科技攻关项目(092102110128);郑州市重大科技专项(112PZDZX030)

摘　　要：根据NCBI网站上萼脊兰AP1-like全长基因序列设计引物,采用RT-PCR法从萼脊兰花瓣中扩增Se AP1-like基因,对扩增产物进行克隆和测序。结果表明,获得的萼脊兰Se AP1-like基因为743 bp,编码247个氨基酸。将Se AP1-like基因插入p CAMBIA1304载体中,经PCR、双酶切鉴定,证实重组表达质粒中含有目的片段,表明成功构建了高效植物表达载体p CAMBIA1304-Se AP1。对Se AP1-like基因的结构域和所表达蛋白质的亲水性进行了分析,并预测了该基因所表达蛋白的二维结构。结果显示,该基因包含MADS-box和K-box保守域,属于MADS-box基因家族,该蛋白分子属于亲水性蛋白。二级结构分析表明,其包含57.49%的α螺旋、6.07%的延伸链、36.44%的不规则折叠。The primers were designed according to the reference Sedirea japonica AP1-like gene from NCBI.The Se AP1-like gene was amplified from the scape of Sedirea japonica petal by RT-PCR.The PCR product was sequenced.The sequencing result showed the target gene was 743 bp encoding 247 putative amino acid residues.The recombinant plasmid p CAMBIA1304-Se AP1 was obtained by identified by inserting the Se AP1-like gene into p CAMBIA1304 vector,and identified by PCR,double digestion.The results showed that the recombinant plasmid p CAMBIA1304-Se AP1 which contained the target gene was constructed successfully.The structural domain of the Se AP1-like gene and the hydrophily of the protein expressed by the Se AP1-like gene were analyzed,and the secondary structure of the protein expressed by the Se AP1-like gene was predicted.The results showed that Se AP1-like gene contained the MADS-box and Kbox domain,belonged to MADS-box super family.This protein expressed by the Se AP1-like gene was hydrophilic and contained 57.49% α-helical domains,6.07% extended strand,36.44% random coil.

关 键 词：萼脊兰 SE AP1-like基因 克隆 载体构建 

分 类 号：S682.31[农业科学—观赏园艺]