Ion Implantation Mediated Glycyrrhiza Genomic DNA Transformation To Yeast

Glycyrrhiza is a important medicinal plant. Glycyrrhizic acid (GA) which is also called Glycyrrhiza saponin or glycyrrhizin, is one of the most important pentacyclic triterpene steroid saponins with biological activity material of the underground part of Glycyrrhiza. Its biosynthesis is controlled by a group of genes. Now we can not construct plasmid vector of target gene for incompletely knowing its genetic background, so there is important theoretical and practical significations to study genetic transformation of Glycyrrhiza genome DNA.In this paper, N+ and Ar+ with 15×1015 ions/cm2 dose were respectively implanted to mediate Glycyrrhiza genome DNA transformation to yeast in the preliminary studies, and 1266 strains of T2 generation recombined yeasts are obtained (772 strains of these by N+ implantation and 494 strains by Ar+ implantation). By taking flavonoids and saponins material as the object products, the results of a qualitative test showed that the rate of yeasts transformation mediated by N+ implantation is lower than that of Ar+ , but its T3 generation of genetic stability higher than that of the latter. This means that the N+ with the biological activity is implanted to mediate Glycyrrhiza genome DNA transformation to yeasts, and its quality sedimentation plays a stimulative role on the genetic stability of recombined yeasts.With TLC chromatography experiments for the metabolic products of recombined yeasts that T4~T8 generations strains, 12 recombined strains with genetic stability that produced glycyrrhizin and (or) glycyrrhetinic acid, 2 recombined strains with genetic stability that produced Glycyrrhiza Flavonoids, and 2 recombined strains produced new red components were obtained. RP-HPLC test results showed that the maximum output of GA of recombined strains with genetic stability was 114.49 mg / L, while that of 18-αand 18-βglycyrrhetinic acid (GAs) were 0.56 mg / L and 0.81 mg / L respectively.The recombined strains are valuable materials to study the key function genes of GA biological synthesis, and to produce GA by microorganism fermentation. It will be a far-reaching impact on study of arid and semi-arid desert medical plants, protection of ecological environment and modernization of traditional Chinese Medicine.