Research Of Hairy Roots Induced And Quality Of Scutellaria Baicalensis Georgi

In this paper, research of hairy roots induced by Agrobacterium rhizogene and cloned β-Glucuronidase gene of Scutellaria baicalensis Georgi, to study the content of baicalin and baicalein and the expression of beta Glucuronidase gene of Scutellaria baicalensis Georgi under different conditions aiming to procide theoretical references for the study of baicalin biosynthetic pathway and metabolic engineering.The hairy root of Scutellaria baicalensis Georgi is successfully induced, and there were two advantages comparing with traditional method. Firstly, the cultived time is reduced, and the agrobacterium on the surface of root is inhibited when using 500mg/L ampicillin sodium sterile distilled water invasion exophyte. Secondly, exophyte is easily survived when adding exogenous hormones into inductive medium.The β-Glucuronidase gene( sbGUS) cDNA from hairy root of Scutellari abaicalensis Georgi. Leafs was cloned by RT-PCR, with GenBank accession number KR364726. The full length cDNA of sbGUS was 1584 bp with an open reading frame(ORF), encoding an unstable protein with 527 amino acids. The bioinformatic analysis show that the sbGUS encoding protein has isoelectric point(pI) of 5.55 and a calculated molecular weight about 58.7248 kD, with a transmembrane regions and signal peptide, has conserved domains of Glycoside hydrolase super family and unintegrated Trans-glycosidase catalytic structure. In the secondary structure, the percentage of alpha helix, Extended strand, β-extended and random coil were 25.62%, 28.84%,13.28% and 32.26%, respectively. The homologous analysis indicates the nucleotide sequence 98.93% similarity and the amino acid sequence 98.29% similarity with Scutellaria baicalensis(BAA97804.1), in the nine positions were different. The expression level of sbGUS was highest in root based on a real-time PCR analysis, followed by flower and stem, and the lowest was in stem. The result will provide a foundation for exploring the molecular function of sbGUS involved in baicalein biosynthesis based on synthetic biology approach in Scutellari abaicalensis plants.By studying the effect of light for the relationship among the content of baicalin and baicalein, the activity of sbGUS and its gene expression, we get the following rules:Firstly, the content of baicalin increased gradually with increasing temperature, and it is indicated that high temperature is benefit to the accumulation of baicalin. Secondly, baiclein of hairy root is easily accumulated in the dark condition, on the contrary, baicalin of hairy root is largely accumulated in light condition. Finally, due to the gene of sbGUS controlled the transformation from baicalin to baicein, in general case, the content of baiclein, the activity of sbGUS enzyme and gene expression levels are positively proportional relationship, however, the content of baicalin is inverse proportion relationship with these elements.