Transformation of filamentous fungi with DNA has been hampered by the low frequencies generally obtained. A variety of methods have been explored, most relying on the enzymatic removal of the cell wall (protoplasting) before DNA treatment (reviewed in (1)). Dithiothreitol (DTT) 4 has been shown to increase the transformation efficiency of Saccharomyces cerevisiae by electroporation (2) and by calcium chloride-induced competency (3) up to 300-fold, and is also employed in some protocols for Candida albicans that result in a 3to 5-fold improvement (2, 4). In order to assist our efforts to perform standard molecular biology experiments on nuclear migration mutants of Aspergillus nidulans that sporulate poorly, we amended a protocol for transformation of A. nidulans protoplasts to include DTT. We describe the modified procedure below and show that we are able to increase transformation efficiency significantly above that obtained without DTT.
The data reported here were obtained using strain GR5 (pyrG89, wA2, pyroA4), but we have also tested AO1 (nudC3, wA2, nicA2, pabaA1, pyrG89, chaA1) and DW61 (snf1364, pyroA4, pyrG89). Media consisted of YGΟ©KCl liquid (0.5% yeast extract (Difco), 2% dextrose (Fisher), 0.6 M KCl (Sigma), trace elements (5) or YAG Ο© KCl plates (YG Ο© KCl plus 1.5% agar (Difco)).