Licorice Beta-as Gene Polymorphism And ¦Â-fragrant Resin Alcohol Synthesis Research

Glycyrrhiza uralensis is wildly used in China.Glycyrrhizin level is one of the most important indexes to estimate the quality of medicinal licorice.Wild resources of Glycyrrhiza are limited, glycyrrhizin content in cultivated Glycyrrhiza can not achieve the standards of 2%, hence, how to improve the glycyrrizin content in cultivated Glycyrrhiza is important. To explore the effects of the polymorphism ofβ-amyrin synthase on the variation of glycyrrhizic acid are of great significance to improve the quality of cultivated licorice and realize sustainable use of Glycyrrhiza resources.Glycyrrhizic acid content in 80 cultivated Glycyrrhiza uralensis were determined by HPLC,according to the very significant level (p<0.0001),80 Glycyrrhiza uralensis in accordance with glycyrrhizic acid will be grouped by SAS9.0.Using RT-PCR strategy to amplification the Open Reading Frame ofβ-amyrin synthase with the template of total RNA extracted from roots of Glycyrrhiza uralensis.Then using DNAman to analyze the relationship between glycyrrhizic acid content and the single nucleotide polymorphism ofβ-amyrin synthase(β-AS) in order to selectβ-AS genotypes which are related to high and low glycyrrhizic acid.The 2289bp fragment in plasmid pMD-19T encodingβ-amyrin synthase was subcloned into the yeast-Escherichia coli shuttle vector pY26,thus an expression recombinant plasmid PY26-β-AS contatining target gene was constructed.The PY-β-AS was introduced into defective mutant INVSc1 of Saccharomyces cerevisiae by LiAc method.β-amyrin synthase activity was expressed under appropriate media and temperature conditions,the level ofβ-amyrin was detected by GC-MS.The main conclusions are as follows:(1)The Open Reading Frame ofβ-AS from Glycyrrhiza uralensis is 2289bp in length encoding one pelypeptide of 762 amino acid(GenBank:FJ627179).The conclusion will provid a foundation for exploring the mechanism of triterpenes biosynthesis.(2)A-T genotype,G/A-C genotype and G-T genotype are associated with the high content of glycyrrhizic acid.(3)The catalytic efficiency ofβ-AS(A-T) genotype is higher thanβ-AS(G-C) genotype and provide a foundation for Licorice molecular breeding.