| Literature DB >> 21590421 |
Gyungsoon Park1, Hildur V Colot, Patrick D Collopy, Svetlana Krystofova, Christopher Crew, Carol Ringelberg, Liubov Litvinkova, Lorena Altamirano, Liande Li, Susan Curilla, Wei Wang, Norma Gorrochotegui-Escalante, Jay C Dunlap, Katherine A Borkovich.
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.Entities:
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Year: 2011 PMID： 21590421 PMCID： PMC3671941 DOI： 10.1007/978-1-61779-040-9_13
Source DB: PubMed Journal: Methods Mol Biol ISSN： 1064-3745