title: High-throughput production of gene replacement mutants in Neurospora crassa.
abstract: The model filamentous fungus Neurospora crassa has been the focus of functional genomics studies for the past several years. A high-throughput gene knockout procedure has been developed and used to generate mutants for more than two-thirds of the ∼10,000 annotated N. crassa genes. Yeast recombinational cloning was incorporated as an efficient procedure to produce all knockout cassettes. N. crassa strains with the Δmus-51 or Δmus-52 deletion mutations were used as transformation recipients in order to reduce the incidence of ectopic integration and increase homologous recombination of knockout cassettes into the genome. A 96-well format was used for many steps of the procedure, including fungal transformation, isolation of homokaryons, and verification of mutants. In addition, development of software programs for primer design and restriction enzyme selection facilitated the high-throughput aspects of the overall protocol.
authors: Department of Plant Pathology and Microbiology, University of California, Riverside, CA, USA.
authors: Gyungsoon
authors: G
authors: Park
authors: Hildur V
authors: HV
authors: Colot
authors: Patrick D
authors: PD
authors: Collopy
authors: Svetlana
authors: S
authors: Krystofova
authors: Christopher
authors: C
authors: Crew
authors: Carol
authors: C
authors: Ringelberg
authors: Liubov
authors: L
authors: Litvinkova
authors: Lorena
authors: L
authors: Altamirano
authors: Liande
authors: L
authors: Li
authors: Susan
authors: S
authors: Curilla
authors: Wei
authors: W
authors: Wang
authors: Norma
authors: N
authors: Gorrochotegui-Escalante
authors: Jay C
authors: JC
authors: Dunlap
authors: Katherine A
authors: KA
authors: Borkovich
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