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The development of a heterologous transformation system for the cellulolytic fungus Trichoderma reesei based on a pyrG-negative mutant strain
Authors:F. Gruber  J. Visser  C. P. Kubicek  L. H. de Graaff
Affiliation:(1) Abt Mikrobielle Biochemie, Institut für Biochemische Technologie und Mikrobiologie, TU Wien, Getreidemarkt 9, A-1060 Wien, Austria;(2) Department of Genetics, Agricultural University, Dreijenlaan 2, NL-6703 HA Wageningen, The Netherlands
Abstract:Summary Six uridine auxotroph mutants of Trichoderma reesei QM 9414 were isolated by resistance to 5-fluoroorotic acid and one strain was identified as OMP-decarboxylase negative (pyr-) by a radiometric enzyme assay. Transformation to uridine prototrophy was achieved with the pyr4 gene of Neurospora crassa (up to 1500 transformants/mgrg) and with pyrA of Aspergillus niger (700–800 transformants/mgrg). In many transformants the PYR+ function seems to be present as extrachromosomal DNA. There is evidence for a correlation between the stability of transformants and integration of the vector in the genome whereas unstable transformants are obtained when autonomous replication of the plasmid occurs.
Keywords:Trichoderma reesei  Transformation  OMP-decarboxylase  Vector integration  Autonomous replication
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