中国现代中药2017，Vol.19Issue(9)：1211-1220,10.DOI:10.13313/j.issn.1673-4890.2017.9.002
滇重楼GAox基因的克隆与生物信息学分析
Cloning and Bioinformatic Analysis of GAox Genes from Paris polyphylla var.yunnanensis
摘要
Abstract
Objective:To clone the full-length cDNA sequences of gibberellic oxidase genes in Paris polyphyllavar.yunnanensiswhich encode the key enzymes involved in GA biosynthetic pathway and analyze their bioinformatic properties.Methods:The sequences of candidate genes in our previous transcriptomic database of Paris polyphyllavar.yunnanensis were used to design the gene-specific PCR primers for RT-PCR cloning of their full-length coding regions.The amplified cDNA fragments were transformed into pMD18-T vector and E.coli DH5α competent cells for sequencing.A series of bioinformatic analysis were carried out,including sequence comparison of homologies,reconstruction of GAox phylogenetic tree,domain search,and 3D structure prediction.Results:Sequencing results showed that the full-length CDS of six PpGAoxgenes were about 1000 bp.Six genes,named as PpGA2ox1,PpGA2ox2,PpGA2ox3,PpGA2ox4,PpGA20ox1 and PpGA3ox1,belonged to three GAox subfamilies,encoding 300-400 amino acids.Their theoretical pI values were about 4-7 and in a positive charge.The proteins of PpGA2ox1,PpGA2ox2 and PpGA2ox4 were unstable and PpGA2ox3,PpGA20ox1 and PpGA3ox1 were stable.However,all of them were hydrophilic,signal peptide and had no transmembrane domain.PpGA2oxl was predicted to be located in the chloroplast,and the rest d in the cytoplasm.PpaGAox proteins had higher homologous sequences with those of other species w and strong domain conservatism.Conclusion:The full-length cDNA sequences of six GAoxwere cloned for the first time from P.polyphylla var.yunnanensis and the bioinformatic analysis were carried out.关键词
滇重楼/GA氧化酶/基因克隆/生物信息学分析Key words
Paris polyphylla var.yunnanensis/GA oxidase/gene cloning/bioinformatic analysis引用本文复制引用
张梦幻,廖登群,孙鹏,李先恩,祁建军..滇重楼GAox基因的克隆与生物信息学分析[J].中国现代中药,2017,19(9):1211-1220,10.基金项目
国家自然科学基金(31471575) （31471575）
