Molecular Cloning, Characterization And Expression Of 1-deoxy-d-xylulose 5-phosphate Reductoisomerase (DXR) Gene From Salvia Miltiorrhiza, Molecular Cloning Of The Promoter Of 1-deoxy-d-xylulose 5-phosphate Synthase I (DXSI) From Salvia Miltiorrhiza

Salvia miltiorrhira, is a kind of species which belongs to family Labiatae, genera Salvia. As a kind of important ancient Chinese medicine, Salvia miltiorrhira is used to treat cardiovascular diseases such as particularly angina pectoris and myocardial infarction. On the basis of the chemical structures, the major bioactive components in S. miltiorrhiza BUNGE can be classified as lipid-soluble diterpenoids and water-soluble polyphenolic compounds. Most of lipophilic constituents are diterpene chinone compounds of the tanshinone type, including tanshinoneâ… , tanshinoneâ…¡A, tanshinone IIB, cryptotanshinone and so on.Terpenes are one kind secondary metabolites which widespread in plants while they are also the largest groups of natural products comprising numerous compounds. All the terpenes are formed through the reactions of IPP (isopentenyl pyrophosphate) and DMAPP (dimethylallyl pyrophosphate), which via two biological reactions, MVA pathway and DXP pathway.The present study describes the cloning and characterization of a cDNA encoding DXR from Salvia miltiorrhiza (designated as SmDXR, GenBank Accession No. FJ476255) by the method of RACE. The cloned full-length cDNA of contained a 1425bp ORF encoding a deduced protein of 475 amino acid residues. Comparative and bioinformatic analyses revealed that SmDXR showed extensive homology with DXRs from other plant species. On the amino acid level, SmDXR was 98% and 91% identical to Mentha x piperita DXR and Antirrhinum majus DXR. According to the phylogenetic tree, SmDXR belongs to the plant group and has the closest relationship with DXR from Lycopersicon esculentum. Tissue expression analysis showed that SmDXR was differentially expressed in various tissues and was found to be higher in leaves followed by roots and stems, indicating SmDXR was considered to be a constitutively expressing gene, which is similar with the most DXRs which have been reported. Expression analysis of SmDXR under induction of SA and MeJA showed that SmDXR can be induced by SA while can not be induced by MeJA. Color complementation assay confirmed SmDXR encoded a functional protein and played an important role in promoting DXP pathway flux.Additionally, the DXSI's five prime untranslated region and promoter sequence of 862bp was cloned for the first time by LA-PCR. The predicting result of PlantCARE website indicates that the sequence has not only some basic elements including TATA-box and CAAT-box but also some stress-induced elements, for example, ABRE (cis-acting element involved in the abscisic acid responsiveness), cis-acting regulatory element involved in the MeJA-responsiveness, CGTCA-motif (cis-acting regulatory element involved in the MeJA-responsiveness), GARE-motif (gibberellin-responsive element), TCA-element (cis-acting element involved in salicylic acid responsiveness) and so on.