The Research Of The Cloning And Expression Analysis For The Rosmarinic Acid Synth Ase Gene In Perilla Frutescens

In this paper, Rosmarinic acid synthase (RAS) gene was firstly cloned from the young leaves of Perilla frutescens which was one of the Labiatae plants by rapid amplification of cDNA ends(RACE) technique and it was named PerRAS(Accession NO. KC355369). Biology information and expression profiling of the novel gene was researched.The primers of PerRAS were designed based on sequences of other RAS genes, such as Salvia miltiorrhiza, Solenoatemon scutellarioides and Melissa officinalis. The full-length of Per RAS cDNA was 1516bp, containing 1299bp open reading frame which encoded a pepitide of 432 amanio acides, and two non-coding regions,72bp 5’UTR and 145bp 3’UTR. The result of BLAST comparision revealed that PerRAS sequence shared 89.76%,87.67%, 87.45% and 79.73% indentity with Melissa officinalis, Salvia miltiorrhiza, Solenoatemon scutellarioides and Lavandula angustifolia, respectively. The amino acid sequence shared 84.13% identity with Melissa officinalis, Salvia, Solenoatemon scutellarioides and Lavandula angustifoliaThe PerRAS sequence was analysised by bioinformatics software. The result showed that the amino acids was hydrophobic; the research of signal peptide revealed that there wasn’t signal peptide Restriction Enzyme cutting site, so the signal peptide didn’t exist. Meanwhile, there wasn’t transmembrane domain. The secondary structure prediction of RAS showed that it contained 19.68% alpha helix,24.30% outspread chains and 56.02% random curly. The three-dimensional model was created for the further research of the senior structure. Phylogenetic tree analysis revealed that each RAS showed different characteristics obviously, denpending on the genera it belonged to. RAS of Perilla frutescens had closer relationship with RASs from Lamiaceae plants than from other plants, and RAS gene was ancient existing early in angiosperms and gymnosperms.Expression profiling analysis of PerRAS was researched by Real-time fluorescent quantitative PCR, β-Actin used as internal standard. The relative expressed quantity of PerRAS gene showed that the position of the highest expression level of PerRAS is roots and the weakest is stems. It revealed that the RAS may exercise the main catalytic effect in roots. Further expression analysis indicated that the Ultraviolet-B radiation(UV-B), H2O2 treatments could down-regulate the PerRAS transcript level and MeJA treatment could up-regulate its transcription level in leaves compared with the control in some degree. It was inferred that PerRAS could assist Perilla frutescens resisting oxidative stress.