Cloning, Expression Analysis And Genetic Transformation Of MicroR828in Salvia Miltiorrhiza

Salvia miltiorrhiza Bunge is one of the most important traditional Chinese herbal medicine. It is labiatae sage and a perennial herb, the root of S. miltiorrhiza (known as Danshen in Chinese) is used for medicine. it have Bitter taste and micro cold. The chief functions were known as the invigoration of circulation of blood, nourishing the blood and tranquilization and so on. It widely distributed in east China, north China, central south and northwest, and it also distributed in part regions of southwest.microRNAs (miRNAs), a class of endogenous small RNA molecules with sizes about20-25nucleotides and a kind of negative regulatory factors, play vital regula-tory roles in eukaryotes. They does not contain open reading frame (ORF). miRNAs regulate gene expression in living beings post-transcriptionally, either by translational inhibition or endonucleolytic cleavage. In plants, miRNAs major in regulating gene expression by endonucleolytic cleavage, and they can regulate plant growth, deve-lopment, morphogenesis, secondary metabolism process and so on. In a word, miRNAs play an important role in plants. In recent years, investigators discover that microR828(miR828) can target MYB genes in some plants, MYB is one of the largest family in plant transcription factors, they can be involved in the regulation of plant secondary metabolic pathways of phenyl propane, the way is one of the main three secondary metabolic pathway in plants. The wild resources of S. miltiorrhiza are in gradually reducing, the use of biotechnology to improve the effective medicinal active ingredients in S. miltiorrhiza is of great importance.Here, we report the cloning of full-length MIR828gene from S. miltiorrhiza using RACE-PCR technology. MIR828is a single-exon gene with595base pairs in length. We examined miR828precursor and mature in four organs of S. miltiorrhiza using the SYBR Green RT-PCR assay. The results showed that the quantity of the precursor and the mature in flower are both relatively higher than in root, stem and leaf. It can establi-sh certain theoretical foundation for the following reserch of miR828function in S.milti-orrhiza.Then we constructed the over-expression vector of miR828and introduced into leaf of S.miltiorrhiza by Agrobacterium tumefaciens-mediated transformation method. Finally, we obtained2transgenic lines, laiding the foundation for further verifying the function of miR828and the use of the genetic engineering means to improve the genetic quality of S.miltiorrhiza.