Increasing polysaccharide yield by over-expression of OsUgp2 in Ganoderma sinense
过量表达OsUgp2基因提高紫芝多糖含量

UDP-glucose pyrophosphorylase (UGPase) plays an important role in the process of polysaccharide synthesis. Two UGPase homologous genes existing in rice genome were designated as OsUgp1 and OsUgp2. In this study, an over-expression vector, in which OsUgp2 gene was driven by a promoter of glyceraldehyde-3-phosphate dehydrogenase from Aspergillus nidulans, was constructed and introduced into Ganoderma sinense via Agribacterium tumefaciens mediated method. A number of hygromycin resistant transformants was obtained. PCR and Southern blot analyses showed that the exogenous OsUgp2 was integrated into the genome of Ganoderma sinense. The result of semi-quantitative RT-PCR indicated that OsUgp2 expressed and the expression level was different among the three independent transformants. The determined UGPase activity of transformants was found increased and its extent was correlated well with the mRNA level. The constant weight and phenol-sulfuric acid assays were utilized to determine the mycelial biomass and polysaccharide content respectively. The results showed that all of biomass, intracellular and extracellular polysaccharides were evidently increased. The positive correlation between UGPase activity and polysaccharide content was revealed by analysis using SPSS13.0 software. The correlation coefficient between UGPase activity and intracellular polysaccharide was the highest at 0.980. It is suggested that the overexpressed OsUgp2 is probably more concerned with intracellular polysaccharide synthesis.