Abstract

Publisher Summary This chapter describes fungal transformation based on the HmB resistance gene of E. coli (hph) and the phleomycin resistance gene of S. hindustanus (ble). To permit selection of transformed cells, markers are used that are capable of complementing a mutation (auxotrophic markers) or that provide a new property to the host cell, such as antibiotic resistance (dominant selectable markers). The latter type of selection has the advantage that mutant strains are not required. This implies that genetically uncharacterized species (among which many pathogenic, and industrially used species) can also be transformed using dominant selectable markers. Two of the markers, which are widely used for fungal transformation, provide resistance against hygromycin B (HmB) and phleomycin (Phle). Hygromycin B is an aminoglycosidic antibiotic that disturbs protein synthesis by interfering with peptidyl-tRNA translocation and causing misreading. Hygromycin B resistance genes, isolated from Streptomyces hygroscopicus 5 and Escherichia coli , encode HmB phosphotransferases that inactivate the antibiotic by phosphorylation. Phleomycin is a metalloglycopeptide antibiotic causing DNA strand scission. Phleomycin resistances genes, isolated from Streptoalloteichus hindustanus and E. coli, encode proteins that inactivate the antibiotic by binding to it.

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