Efficient cloning of genes of Neurospora crassa

We have constructed a genomic library of Neurospora crassa DNA in a cosmid vector that contains the dominant selectable marker for benomyl resistance. The library is arranged to permit the rapid cloning of Neurospora genes by either sib-selection or colony-hybridization protocols. Detailed procedures for the uses of the library are described. By use of these procedures, a modest number of unrelated genes have been isolated. The cloning of trp-3, the structural gene for the multifunctional enzyme tryptophan synthetase (tryptophan synthase, EC 4.2.1.20), is reported in detail; its identity was verified by restriction fragment length polymorphism mapping. The strategies described in this paper should be of use in the cloning of any gene of Neurospora, as well as genes of other lower eukaryotes.