Full-length CDNA Cloning Of Strictosidine Synthase In Rauvolfia Yunnanensis Tsiang

Rauvolfia yunnanensis Tsiang is an important medicinal plant, which contains a variety of terpenoid indole alkaloids (terpenoid indole alkaloids, TIAs). It's the bulk drugs for the treatment of diseases such as hypertension, sexual dysfunction and cancer. Strictosidine synthase work as a catalytic. It catalyze tryptamine and secologanin condensation to3α(S)-Strictosidine. STR is the key intermediate of Rauvolfia yunnanensis Tsiang terpenoid indole alkaloids sencondary metabolism'pathway. It haven't been reported from the Rauvolfia yunnanensis Tsiang. So the study on STR of Rauvolfia yunnanensis Tsiang is meaningful for the improve the production of TIAs. The major findings of this paper are as follows:1) Full-length cDNA cloning of STR gene from Rauvolfia yunnanensis Tsiang. Using the fresh leaf of Rauvolfia yunnanensis Tsiang as the material, to extract total RNA, and transcript it reversed to cDNA as a template. Based on the nucleotide sequence fragments in NCBI, through a professional software, Primer Premier5.0, to designe specific primers. Then amplified by PCR, the fragment was cloned into the vector pMD18-T and then transformed into Escherichia. Coli DH5a and its sequence was obtained by sequencing recombinant plasmid DNA extracted from the transformed strain. After analyzing by DNAMAN,the full-length of a1038bp STR gene cDNA cloning from Rauvolfia yunnanensis Tsiang were obtained.2) Bioinformatic analysis of STR from Rauvolfia yunnanensis Tsiang. We use the DNAMAN to analysis the sequences of STR genes, the1038bp STR cDNA sequence has a1037bp ORF coding345amino acids. The homology percentage of the STR from Rauvolfia yunnanensis Tsiang with other Rauvolfia species in Genbank reach up to99%,and homology percentage with Catharanthus roseus reach up to80%. The characteristic of physical chemistry and the second structure of the deduced amino acid sequences were predicted and analyzed by Bio-softwares, and the results for STR are:pl value is5.20; molecular weights is38.4kDa; the instability index is34.76; the aliphatic index is86.70; average hydrophilia is 0.132.And27amino acids constructed a signal peptide that attend excrete at its N-end.The prediction of its secondary structure shows that extend strand and random coil is the most quantities in STR protein and α-spiral and β-outer corner distribute in the whole protein.