苦参查尔酮异构酶的基因克隆与表达分析

Cloning and Expression Analysis of Chalcone Isomerase from Sophora flavescens

目的为明晰苦参查尔酮异构酶(sfCHI)的结构基因信息及其在苦参黄酮类化合物生源路径中的表达调控机理。方法本实验以苦参为研究对象,基于其转录组学数据,克隆sfCHI基因并针对该cDNA序列展开生物信息学分析,采用半定量PCR方法检测安徽产苦参不同组织中sfCHI基因表达情况。结果结果显示苦参sfCHI基因的cDNA序列长度为1265 bp,含有630 bp的开放式阅读框(ORF),编码209个氨基酸;序列分析表明sfCHI与狭叶羽扇豆查尔酮异构酶基因相似性最高,达89.95%;进化树分析该基因与大豆、野大豆等植物的亲缘关系较近;半定量PCR结果显示sfCHI基因在苦参根中表达量最低,在叶、叶柄与茎中的表达未见明显差异。结论本研究通过对苦参查尔酮异构酶的基因克隆与不同组织间的表达分析,初步明确了该基因的序列信息与理化特征,这对进一步研究苦参黄酮类物质生物合成机制将起到积极的推动作用。

Objective In order to clarify the structural gene information of Sophora flavescens chalcone isomerase(sfCHI)and its expression pattern among distinct organs.Methods This experiment took advantage of S.flavescens deep transcriptomic data to clone sfCHI gene and conducted bioinformatics analysis on the cDNA sequence.Semi-quantitative PCR was used to detect the expression of sfCHI gene in different organs of S.flavescens.Results The results showed that the cDNA sequence of S.flavescens CHI gene was 1265 bp in length and contained a 630 bp open reading frame(ORF)encoding 209 amino acids.Sequence analysis showed that the similarity between sfCHI and Lupinus angustifolius CHI gene was the highest,up to 89.95%.Phylogenetic tree resulted from 16 selected CHI genes showed that sfCHI and other related genes from Leguminosae were in the same group.Semi-quantitative PCR results showed that sfCHI gene expression in the root of S.flavescens is the lowest,while expression in leaves,petioles and stems did not show obvious difference.Conclusion In this study,the sequence information and physicochemical characteristics of the gene were firstly clarified by gene cloning of the chalcone isomerase gene and the expression analysis of different tissues,which will promote the study of biosynthetic mechanism of flavonoids in S.flavescens.