农业生物技术学报2024，Vol.32Issue(3)：533-545,13.DOI:10.3969/j.issn.1674-7968.2024.03.004
钩吻GeCYP85A1基因的克隆与表达分析
Cloning and Expression Analysis of the GeCYP85A1 Gene in Gelsemium elegans
摘要
Abstract
Gelsemium elegans is a valuable traditional Chinese herb in China.The whole plant is highly toxic,but has anti-tumour and analgesic effects,as well as fattening effects on pigs and other livestock,and its main active constituents are terpenoid indole alkaloids(TIAs).Among them,the most representative ingredient is gelsenicine,which has the strongest toxicity and highest activity.In this study,209 genes related to cytochrome P450(CYP450)were screened based on the genomic and transcriptomic data of G.elegans,combined with metabolomic data,an important candidate gene GeCYP85A1(GenBank No.OQ653843)related to the biosynthesis pathway of Oryza japonica was screened by"genome+expression profile+metabolome".The full-length cloning of this gene was performed by PCR,and its bioinformatics analysis was carried out.The expression pattern of the GeCYP85A1 gene in different tissues of G.elegans was analysed by qPCR.The results showed that the GeCYP85A1 gene was 1 392 bp long,encoding 463 amino acids,with a theoretical relative molecular mass of 53.390 06 kD and a theoretical isoelectric point of 9.07.It was a non-secretory protein with a subcellular localization in chloroplasts,a transmembrane region and no signal peptide.It was similar to Coffea arabica,Coffea eugenioides,Erythranthe guttata,Nicotiana tabacum,Olea europaea subsp.europaea,Phtheirospermum juttata,Salvia miltiorrhiza,Sesamum indicum,and Vitis riparia,and the amino acid sequences of the related CYP85A1 of the above 9 species were highly similar and belonged to a subfamily of the CYP450 family.By qPCR,the expression pattern of GeCYP85A1 in different parts of the plant was found to be consistent with the trend of the relative content of gelsenicine in different parts of the plant,which were root＞flower＞stem＞leaf.This study provides a reference for further elucidation of the biosynthetic pathway for the TIAs-like alkaloids in G.elegans.关键词
钩吻/GeCYP85A1/生物信息学分析/表达分析Key words
Gelsemium elegans/GeCYP85A1/Bioinformatics analysis/Expression analysis分类
农业科技引用本文复制引用
章瑶,陈文强,潘丽梅,穆德添,周宇,林孝东,刘兆颖,唐其..钩吻GeCYP85A1基因的克隆与表达分析[J].农业生物技术学报,2024,32(3):533-545,13.基金项目
湖南省科技计划项目(2022NK2004 （）
2017ZK3133) （）
湖南省教育厅项目(21A0138 （）
HNJG-2020-0327) （）
湖南农业大学研究生科研创新项目(2022XC062) （2022XC062）
