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High efficient homologous integration via tandem exo-beta-1,3-glucanase genes in the common mushroom, Agaricus bisporus
- Rhee, M.D. van de, Mendes, O., Werten, M.W.T., Huizing, H.J., Mooibroek, H.
- Current genetics 1996 v.30 no.2 pp. 166-173
- genetic transformation, glycosidases, messenger RNA, restriction mapping, structural genes, Agaricus bisporus, open reading frames, genetic code, plasmid vectors, amino acid sequences, homologous recombination, gene expression, fruiting bodies, introns, mycelium
- Homologous integration was studied in the common mushroom, Agaricus bisporus, using a plasmid (pHAG3-1) carrying the hygromycin-resistance gene and a 3.2-kb genomic fragment from A. bisporus. Homologous integration was found in 30-60% of the transformants obtained with pHAG3-1 linearized at three different positions within the homologous sequence, generating either blunt, 5'- or 3'-protruding ends. The genomic fragment was found to contain two homologous open reading frames in tandem, which showed 60% similarity to exo-beta-1,3-glucanases from Saccharomyces cerevisiae and Candida albicans. The level of the corresponding mRNA is low in the vegetative mycelium and relatively high in fruiting bodies. In the vegetative mycelium of a transformant with tandemly integrated pHAG3-1 plasmids at the homologous position, exoglucanase mRNA was strongly increased without any apparent effect on growth rate or morphology.