A full length cDNA of 4-(cytidine 5'-diphospho)-2-C-methyl-D-erythritol kinase cloning and analysis of introduced gene expression in Salvia miltiorrhiza / 药学学报

ABSTRACT

This paper firstly introduced the acquired full length cDNA of 4-(cytidine 5'-diphospho)-2-C-methyl-D-erythritol kinase from hairy roots of Salvia miltiorrhiza (Abbr SmCMK, GenBank number EF534309). Results of KEGG analysis showed that SmCMK was belong to the upstream of nonemevalonate pathway, the only one kinase of the pathway. The full-length cDNA was deduced as encoding 4-(cytidine 5'-diphospho)-2-C-methylerythritol kinase (designated as SmCMK), and the sequence had a 1493 bp including 5' UTR 71 bp and 3' UTR 232 bp, an open reading frame (ORF) encoding a protein of 396 amino acid residues. The deduced protein had isoelectric point (pI) of 6.78 and a calculated molecular weight about 43 kDa, similar to cloned diterpene of CMK from other species of plants such as Mentha piperita and Lycopersicon esculentum reported previously. Real time PCR results indicated that elicitors of MJ stimulated the increase of mRNA expression of SmCMK. At the same time, results of high performance liquid chromatography (HPLC), used to examine the accumulation of diterpenoid tanshinones in hairy roots, showed that the contents of diterpenoid tanshinones in hairy root of Salvia miltiorrhiza were increased dramatically after treated with methyl jasmonate (MJ). This result showed a positive correlation between the levels of mRNA expression and tanshinones accumulation in Salvia miltiorrhiza stimulated by MJ. It proved primarily that the increased expression level of mRNA of SmCMK helps to enhance tanshinones' accumulation, which will be the basis for further study on the mechanism of gene regulation of secondary metabolism of tanshinones.