Characterization of glyceraldehyde-3-phosphate dehydrogenase gene RtGPD1 and development of genetic transformation method by dominant selection in oleaginous yeast Rhodosporidium toruloides

Cloning of glyceraldehyde-3-phosphate dehydrogenase genes from the basidiomycete mushroom Pleurotus ostreatus and analysis of their expression during fruit-body development

“…In L. edodes, the GPD gene is expressed constitutively during each stage of fruit-body development and in the two parts of the mature fruit body, the gills and cap (Hirano et al 1999). However, GPD gene expression has been reported to be regulated under specific metabolic and environmental conditions in S. cerevisiae (McAlister and Holland 1985;Boucherié et al 1995), L. sajor-caju (Jeong et al 2000), and R. toruloides (Liu et al 2013), and during cell growth and differentiation in T. harzianum (Puyesky et al 1997), P. brasiliensis (Barbosa et al 2004), and P. crystallinus (Kubo 2012), although the reason for regulation is not fully understood. Accordingly, our results suggest that the expression of PoGPD1 and PoGPD2 is developmentally regulated in P. ostreatus strain H1, PoGPD1 and PoGPD2 proteins may serve to supply energy and substrates through glycolysis and gluconeogenesis preferentially in the mycelial and primordial stages and in the base as compared to other parts of mature fruit bodies.…”

“…However, previous studies have demonstrated that the expression of GPD genes is regulated by several cellular conditions in some fungi. For instance, under conditions of environmental stress such as high salt content and cold, GPD gene expression is up-regulated in Lentinus sajor-caju (Jeong et al 2000) and Rhodosporidium toruloides (Liu et al 2013). During cellular growth and differentiation, GPD gene expression is induced during the transition from mycelium to yeast in Paracoccidioides brasiliensis (Barbosa et al 2004), while GPD gene expression is repressed during conidiation and mycoparasitism in Trichoderma harzianum (Puyesky et al 1997).…”

Agrobacterium tumefaciens- Mediated Transformation of Pucciniomycotina Red Yeasts

“…In developing these vectors, and comparing their ability to work in different species, a high degree of specifi city was found between the source of gene markers and the species to transform. This specifi city seems to be due to intrinsic features of the red yeast itself, the percentage of G + C content in its genome, and/or codon usage (Abbott et al 2013, Liu et al 2013 ). Transformation of red yeasts became successful upon utilization 1 Six different binary vectors are available for transformation of Pucciniomycotina red yeasts.…”

“…Recently, protocols for A. tumefaciens -mediated transformation (AMT) of red yeasts became available (e.g. Ianiri et al 2011, Abbott et al 2013, Liu et al 2013 ). An alternative approach is the use of biolistic delivery of DNA into these species (Ianiri et al 2011 ).…”

“…For selection and maintenance of these vectors in bacteria, 50 μg/ml of kanamycin is added to Luria-Bertani medium. The vector pRH2031 features a codon-optimized enhanced green fl uorescent protein gene (Rt GFP ) and hygromycin phosphotransferase gene ( hpt -3 ) placed under the control of the promoter and terminator of the glyceraldehyde-3-phosphate dehydrogenase gene ( GPD1 ) of Rhodosporidium toruloides (see Liu et al 2013 for details). For selection and maintenance of the vectors in bacteria, 50 μg/ml of rifampicin and 50 μg/ml of spectinomycin are added to the medium.…”