穿心莲甲羟戊酸5-焦磷酸脱羧酶基因的克隆与表达

Cloning and tissue expression of mevalonate disphosphate decarboxylase genes from Andrographis paniculata

穿心莲内酯是穿心莲的主要药效成分,广泛用于抗炎。为了对它的生物合成进行遗传调控,提高内酯类成分的合成量,作者在转录组数据分析的基础上,获得了穿心莲内酯生物合成途径中甲羟戊酸5-焦磷酸脱羧酶基因的3个克隆,包含的ORF均长1 260 bp,分别在4个位点具有单碱基的差异。它们编码由419个残基组成的氨基酸序列;保守结构域中均具有11个高度保守的氨基酸,分别决定催化反应的特异性和活性;N端不含有质体定位的信号肽;与丹参的MVD蛋白(Gen Bank号AEZ55675.1)一致性较高。在穿心莲茎和叶片中,MVD基因的表达量分别在花蕾期和初花期最高,但各时期表达量的倍数差异不大。该研究获得的MVD基因,为后续进行详细的功能解析,并进一步应用于穿心莲内酯的遗传调控,奠定了基础。

Andrographolide is a main bioactive substance in Andrographis paniculata,and extensively used in anti-inflammatory drugs. In order to increase andrographolide production in plant,three 1260 bp ORFs encoding mevalonate disphosphate decarboxylases with 419 amino acids were cloned from A. paniculata by RACE method and analyzed by bioinformatic software. Their tissue expression patterns were predicted by real time PCR. Eleven conserved amino acid residues determining specificity and activity of these MVDs were predicted in these amino acid sequences,but no plastid targeted signal peptides were detected. These MVDs have high similarities with the MVD protein（ Gen Bank number： AEZ55675. 1） from Salvia miltiorrhiza. In stems and leaves,expression levels of these MVD genes were constant,and reached the highest level at bud stage and the beginning of flowering. The MVD genes we have cloned from A.paniculata could be used in genetic engineering of andrographolide biosynthsis pathway in future.