ABSTRACT
<p><b>OBJECTIVE</b>To analyze the correlation between content of glycyrrhizic acid and the single nucleotide polymorphism of beta-amyrin synthase (bAS) in Glycyrrhiza uralensis.</p><p><b>METHOD</b>glycyrrhizic acid content in 80 samples of the cultivated G. uralensis were determined by HPLC; According to the very significant level (P < 0.000 1), 80 samples in accordance with glycyrrhizic acid will be grouped by SAS 9.0; Using RT-PCR strategy to amplification the Open Reading Frame of beta-amyrin synthase with the template of total RNA extracted from roots of G. uralensis and then using DNAman to analyze the relationship between glycyrrhizic acid content and the single nucleotide polymorphism of beta-amyrin synthase (bAS).</p><p><b>RESULT</b>There exited two mutation sites 94 bp and 254 bp, G/A conversion occurred at 94 bp site, which belonged to a missense mutation. G/A conversion led to the corresponding amino acid conversion (Gly --> Asp); C/T conversion occurred at 254 bp site, which belonged to a synonymous mutation. According to sequence variation, the samples were divided into four genotypes: G-T genotype, A-T genotype, G/A-C genotype and G-T genotype.</p><p><b>CONCLUSION</b>A-T genotype, G/A-C genotype and G-T genotype are correlated with the high content of glycyrrhizic acid.</p>
Subject(s)
Genotype , Glycyrrhiza uralensis , Genetics , Metabolism , Glycyrrhizic Acid , Metabolism , Intramolecular Transferases , Genetics , Polymorphism, Single Nucleotide , Reproducibility of ResultsABSTRACT
<p><b>OBJECTIVE</b>To analyze heterologous expression in Saccharomyces cerevisiae of two genotypes: beta-AS (A-T) genotype which is related to high content of glycyrrhizic acid and beta-AS(G-C) genotype which is related to low content of glycyrrhizic acid, and compare two different genotypes on the impact of beta-amyrin production in order to provide a foundation for licorice molecular breeding.</p><p><b>METHOD</b>The 2 289 bp fragment in plasmid pMD-19T encoding beta-amyrin synthase was subcloned into the yeast-Escherichia coli shuttle vector pY26, thus an expression recombinant plasmid PY-beta-AS containing target gene was constructed. The PY-beta-AS was introduced into defective mutant INVSc1 of S. cerevisiae by LiAc method, after induced by IPTG, the content of beta-amyrin was determined by GC-MS.</p><p><b>RESULT</b>GC-MS analysis demonstrates that the an occurring peak corresponding to beta-amyrin standards was detected with the same retention time, which is absent in the cell transform with empty vector. Results showed the peak was beta-amyrin and the percentage of beta-amyrin in two genotypes: beta-AS (A-T) genotype and beta-AS (G-C) genotype were 19.08% and 1.40%, respectively. Thus the beta-amyrin synthase exhibited the activity of catalyzing 2, 3-oxidosqualene to beta-amyrin.</p><p><b>CONCLUSION</b>The catalytic efficiency of beta-AS(A-T) genotype is higher than that of beta-AS(G-C) genotype, which can lay the foundation for licorice molecular breeding.</p>
Subject(s)
Catalysis , Cloning, Molecular , Genotype , Glycyrrhiza uralensis , Chemistry , Genetics , Intramolecular Transferases , Chemistry , Genetics , Metabolism , Plant Proteins , Chemistry , Genetics , Metabolism , Polymorphism, Genetic , Recombinant Proteins , Chemistry , Genetics , Metabolism , Saccharomyces cerevisiae , GeneticsABSTRACT
<p><b>OBJECTIVE</b>To clone and sequence the open reading frame of beta-amyrin synthase (bAS) from Glycyrrhiza uralensis.</p><p><b>METHOD</b>The primers were designed according to the cDNA sequence of beta-amyrin synthase from G. glabra reported by Hiroaki HAYASHI, and the open reading frame of beta-amyrin synthase was cloned by RT-PCR strategy with the template of total RNA extracted from roots of G. uralensis.</p><p><b>RESULT</b>The GubAS (GenBank Accession number: FJ627179) was 2 289 bp in length encoding one pelypeptide of 762 amino acid. Deduced amino acid sequence had 99%, 92%, 90%, 90% and 89% homology to the amino acid sequence of G. glabra, Lotus japonicus, Pisum sativum, Medicago truncatula, Glycine max, respectively.</p><p><b>CONCLUSION</b>The open reading frame of bAS from G. uralensis is cloned and reported for the first time. The conclusion will provide a foundation for exploring the mechanism of triterpenes biosynthesis.</p>
Subject(s)
Cloning, Molecular , Glycyrrhiza uralensis , Classification , Genetics , Intramolecular Transferases , Genetics , Metabolism , Molecular Sequence Data , Open Reading Frames , Phylogeny , Plant Proteins , Genetics , Metabolism , Plants , Classification , GeneticsABSTRACT
Objective:Stephania tetrandra is widely used in traditional Chinese medication.In present study,we studied the resource status of it in Anhui and Jiangxi Province.Methods:We inspected relevant documents to confirm which counties we would investigate;and then,we interviewed many relative local people and carried out sample-plot survey.Results:The resource of wild Stephania tetrandra in the two provinces were both seriously damaged.Conclusion:Large-scale promotion of artificial cultivation was required to ensure the supply and to realize the reasonable and sustainable utility of this medicinal material.