农业生物技术学报2025，Vol.33Issue(6)：1246-1258,13.DOI:10.3969/j.issn.1674-7968.2025.06.006
七叶一枝花糖基转移酶基因PpUGT2的克隆及原核表达
Cloning and Prokaryotic Expression of the UDP-glycosyltransferase Gene PpUGT2 from Paris polyphylla var.chinensis
摘要
Abstract
Saponins are important active components of Paris polyphylla chinensis,but their biosynthetic pathway has not been fully elucidated.Research suggests that UPD-glycosyltransferases(UGTs)are key enzymes in the downstream biosynthetic pathway of polyphyllin.This study based on the correlation analysis between the transcriptome of P.polyphylla var.chinensis and the total saponins content,a glycosyltransferase candidate gene PpUGT2(GenBank No.pq588702)was screened out,which might be involved in the biosynthesis of polyphyllin.The PpUGT2 gene was cloned using PCR technology,and an expression vector was constructed for prokaryotic expression.Expression was induced using IPTG,followed by purification of the recombinant protein.Concurrently,qPCR was employed to analyze the expression patterns of PpUGT2 across different parts of P.polyphylla var.chinensis.The results indicated that PpUGT2 had a total length of 1 773 bp,encoded a protein consisting of 590 amino acids with a relative molecular weight of 64.6 kD.This hydrophilic protein was unstable and exhibited good lipid solubility.SDS-PAGE analysis confirmed the expression of the PpUGT2 recombinant protein at approximately 64.6 kD.Furthermore,qPCR results demonstrated expression of the PpUGT2 gene in roots,stems,leaves,and young fruits of P.polyphylla var.chinensis,with a distinct expression hierarchy of roots＞stems＞leaves＞young fruits,suggesting a predominant role of this gene in root tissues.This study enriches the study of UGT in P.polyphylla var.chinensis and provides a theoretical basis for further analysis of the biosynthesis pathway of polyphyllin.关键词
七叶一枝花/尿苷二磷酸糖基转移酶(UGTs)/生物信息学/原核表达/qPCRKey words
Paris polyphylla var.chinensis/UDP-glycosyltransferases(UGTs)/Bioinformatics analysis/Prokaryotic expression/qPCR分类
农业科学引用本文复制引用
王丽娅,王靖涵,周慧敏,章瑶,陈文强,鲁志国,朱丽娜,唐其..七叶一枝花糖基转移酶基因PpUGT2的克隆及原核表达[J].农业生物技术学报,2025,33(6):1246-1258,13.基金项目
湖南省教育厅项目(22C0106) （22C0106）
湖南农业大学创新训练项目(XCX2023160) （XCX2023160）
