Abstract
Pleurotus eryngii was transformed using a polyethylene glycol-mediated method. A plasmid, pEPUGH, containing a reporter gene (enhanced green fluorescent protein gene, egfp) and a positive selectable marker gene (hygromycin phosphotransferase gene, hph) was constructed. The fused egfp-hph gene was placed under the control of the strong and constitutive native gpd promoter from P. eryngii. The recombinant plasmid was used to transform of P. eryngii protoplasts. Successful transformation was demonstrated by molecular analyses. Moreover, the mycelia of the transformants showed green epipolic dispersion on fluorescence microscopy. About 90–210 transformants were produced per μg plasmid DNA per 107 viable protoplasts.
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References
Brandfass C, Karlovsky P (2008) Upscaled CTAB-based DNA extraction and real-time PCR assays for Fusarium culmorum and F. graminearum DNA in plant material with reduced sampling error. Int J Mol Sci 9(11):2306–2321
Burns C, Gregory KE, Kirby M, Cheung MK, Riquelme M, Elliott TJ, Challen MP, Bailey A, Foster GD (2005) Efficient GFP expression in the mushrooms Agaricus bisporus and Coprinus cinereus requires introns. Fungal Genet Biol 42(3):191–199
Ding Y, Liang S, Lei JH, Chen LG, Kothe E, Ma AM (2011) Agrobacterium tumefaciens mediated fused egfp-hph gene expression under the control of gpd promoter in Pleurotus ostreatus. Microbiol Res 166(4):314–322
Gomez-Toribio V, Garcia-Martin AB, Martinez MJ, Martinez AT, Guillen F (2009) Induction of extracellular hydroxyl radical production by white-rot fungi through quinone redox cycling. Appl Environ Microbiol 75(12):3944–3953
Hirano T, Sato T, Yaegashi K, Enei H (2000) Efficient transformation of the edible basidiomycete Lentinus edodes with a vector using a glyceraldehyde-3-phosphate dehydrogenase promoter to hygromycin B resistance. Mol Gen Genet 263(6):1047–1052
Irie T, Honda Y, Hirano T, Sato T, Enei H, Watanabe T, Kuwahara M (2001) Stable transformation of Pleurotus ostreatus to hygromycin B resistance using Lentinus edodes GPD expression signals. Appl Microbiol Biot 56(5–6):707–709
Irie T, Sato T, Saito K, Honda Y, Watanabe T, Kuwahara M, Enei H (2003) Construction of a homologous selectable marker gene for Lentinula edodes transformation. Biosci Biotech Biochem 67(9):2006–2009
Kim S, Song J, Choi HT (2004) Genetic transformation and mutant isolation in Ganoderma lucidum by restriction enzyme-mediated integration. FEMS Microbiol Lett 233(2):201–204
Kim S, Sapkota K, Choi BS, Kim SJ (2010) Expression of human growth hormone gene in Pleurotus eryngii. Cent Eur J Biol 5(6):791–799
Kuo CY, Chou SY, Huang CT (2004) Cloning of glyceraldehyde-3-phosphate dehydrogenase gene and use of the gpd promoter for transformation in Flammulina velutipes. Appl Microbiol Biot 65(5):593–599
Li G, Wang Q, Liu QY, Li BJ (2004) Establishment of a transformation system of Ganoderma lucidum using PEG method. Mycosystema 23:255–261
Li G, Li RX, Liu QY, Wang Q, Chen M, Li BJ (2006) A highly efficient polyethylene glycol-mediated transformation method for mushrooms. FEMS Microbiol Lett 256(2):203–208
Noh W, Kim SW, Dong-Won B, Kim JY, Ro HS (2010) Genetic introduction of foreign genes to Pleurotus eryngii by restriction enzyme-mediated integration. J Microbiol 48(2):253–256
Ohga S (2000) Influence of wood species on the sawdust-based cultivation of Pleurotus abalonus and Pleurotus eryngii. J Wood Sci 46(2):175–179
Acknowledgement
We thank Jason Karakehian and the reviewer for the helpful comments and improvement of the text. This work was supported by the Grant of Beijing Nova Program (Grant No. 2011053) and the Beijing Municipal Committee of CPC Organization Department (Grant No. 2011A002020000005).
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Yin, Y., Liu, Y., Jin, H. et al. Polyethylene glycol-mediated transformation of fused egfp-hph gene under the control of gpd promoter in Pleurotus eryngii . Biotechnol Lett 34, 1895–1900 (2012). https://doi.org/10.1007/s10529-012-0985-5
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DOI: https://doi.org/10.1007/s10529-012-0985-5