Efficient cloning of genes of Neurospora crassa.
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Journal: 2010/June - Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424
PUBMED: 16593723
Abstract:
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.
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Proc Natl Acad Sci U S A 83(13): 4869-4873
PMID: 16593723

Efficient cloning of genes of <em>Neurospora crassa</em>

Abstract

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.

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Selected References

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  • Krumlauf R, Marzluf GA. Characterization of the sequence complexity and organization of the Neurospora crassa genome. Biochemistry. 1979 Aug 21;18(17):3705–3713. [PubMed] [Google Scholar]
  • Perkins DD, Radford A, Newmeyer D, Björkman M. Chromosomal loci of Neurospora crassa. Microbiol Rev. 1982 Dec;46(4):426–570.[PMC free article] [PubMed] [Google Scholar]
  • Case ME, Schweizer M, Kushner SR, Giles NH. Efficient transformation of Neurospora crassa by utilizing hybrid plasmid DNA. Proc Natl Acad Sci U S A. 1979 Oct;76(10):5259–5263.[PMC free article] [PubMed] [Google Scholar]
  • Akins RA, Lambowitz AM. General method for cloning Neurospora crassa nuclear genes by complementation of mutants. Mol Cell Biol. 1985 Sep;5(9):2272–2278.[PMC free article] [PubMed] [Google Scholar]
  • Vapnek D, Hautala JA, Jacobson JW, Giles NH, Kushner SR. Expression in Escherichia coli K-12 of the structural gene for catabolic dehydroquinase of Neurospora crassa. Proc Natl Acad Sci U S A. 1977 Aug;74(8):3508–3512.[PMC free article] [PubMed] [Google Scholar]
  • Schechtman MG, Yanofsky C. Structure of the trifunctional trp-1 gene from Neurospora crassa and its aberrant expression in Escherichia coli. J Mol Appl Genet. 1983;2(1):83–99. [PubMed] [Google Scholar]
  • Buxton FP, Radford A. Cloning of the structural gene for orotidine 5'-phosphate carboxylase of Neurospora crassa by expression in Escherichia coli. Mol Gen Genet. 1983;190(3):403–405. [PubMed] [Google Scholar]
  • Legerton TL, Yanofsky C. Cloning and characterization of the multifunctional his-3 gene of Neurospora crassa. Gene. 1985;39(2-3):129–140. [PubMed] [Google Scholar]
  • Arends H, Sebald W. Nucleotide sequence of the cloned mRNA and gene of the ADP/ATP carrier from Neurospora crassa. EMBO J. 1984 Feb;3(2):377–382.[PMC free article] [PubMed] [Google Scholar]
  • Berlin V, Yanofsky C. Isolation and characterization of genes differentially expressed during conidiation of Neurospora crassa. Mol Cell Biol. 1985 Apr;5(4):849–855.[PMC free article] [PubMed] [Google Scholar]
  • Münger K, Germann UA, Lerch K. Isolation and structural organization of the Neurospora crassa copper metallothionein gene. EMBO J. 1985 Oct;4(10):2665–2668.[PMC free article] [PubMed] [Google Scholar]
  • Paietta JV, Marzluf GA. Gene disruption by transformation in Neurospora crassa. Mol Cell Biol. 1985 Jul;5(7):1554–1559.[PMC free article] [PubMed] [Google Scholar]
  • Dhawale SS, Marzluf GA. Transformation of Neurospora crassa with circular and linear DNA and analysis of the fate of the transforming DNA. Curr Genet. 1985;10(3):205–212. [PubMed] [Google Scholar]
  • Stohl LL, Lambowitz AM. Construction of a shuttle vector for the filamentous fungus Neurospora crassa. Proc Natl Acad Sci U S A. 1983 Feb;80(4):1058–1062.[PMC free article] [PubMed] [Google Scholar]
  • Stohl LL, Akins RA, Lambowitz AM. Characterization of deletion derivatives of an autonomously replicating Neurospora plasmid. Nucleic Acids Res. 1984 Aug 10;12(15):6169–6178.[PMC free article] [PubMed] [Google Scholar]
  • Grant DM, Lambowitz AM, Rambosek JA, Kinsey JA. Transformation of Neurospora crassa with recombinant plasmids containing the cloned glutamate dehydrogenase (am) gene: evidence for autonomous replication of the transforming plasmid. Mol Cell Biol. 1984 Oct;4(10):2041–2051.[PMC free article] [PubMed] [Google Scholar]
  • Paietta J, Marzluf GA. Plasmid recovery from transformants and the isolation of chromosomal DNA segments improving plasmid replication in Neurospora crassa. Curr Genet. 1985;9(5):383–388. [PubMed] [Google Scholar]
  • Hanahan D. Studies on transformation of Escherichia coli with plasmids. J Mol Biol. 1983 Jun 5;166(4):557–580. [PubMed] [Google Scholar]
  • Ish-Horowicz D, Burke JF. Rapid and efficient cosmid cloning. Nucleic Acids Res. 1981 Jul 10;9(13):2989–2998.[PMC free article] [PubMed] [Google Scholar]
  • Chia W, Scott MR, Rigby PW. The construction of cosmid libraries of eukaryotic DNA using the Homer series of vectors. Nucleic Acids Res. 1982 Apr 24;10(8):2503–2520.[PMC free article] [PubMed] [Google Scholar]
  • Hohn B. In vitro packaging of lambda and cosmid DNA. Methods Enzymol. 1979;68:299–309. [PubMed] [Google Scholar]
  • Rigby PW, Dieckmann M, Rhodes C, Berg P. Labeling deoxyribonucleic acid to high specific activity in vitro by nick translation with DNA polymerase I. J Mol Biol. 1977 Jun 15;113(1):237–251. [PubMed] [Google Scholar]
  • Krumlauf R, Marzluf GA. Genome organization and characterization of the repetitive and inverted repeat DNA sequences in Neurospora crassa. J Biol Chem. 1980 Feb 10;255(3):1138–1145. [PubMed] [Google Scholar]
  • Russell PJ, Wagner S, Rodland KD, Feinbaum RL, Russell JP, Bret-Harte MS, Free SJ, Metzenberg RL. Organization of the ribosomal ribonucleic acid genes in various wild-type strains and wild-collected strains of Neurospora. Mol Gen Genet. 1984;196(2):275–282. [PubMed] [Google Scholar]
  • Clarke L, Carbon J. A colony bank containing synthetic Col El hybrid plasmids representative of the entire E. coli genome. Cell. 1976 Sep;9(1):91–99. [PubMed] [Google Scholar]
  • DEMOSS JA. Studies on the mechanism of the tryptophan synthetase reaction. Biochim Biophys Acta. 1962 Aug 13;62:279–293. [PubMed] [Google Scholar]
  • Miles EW. Tryptophan synthase: structure, function, and subunit interaction. Adv Enzymol Relat Areas Mol Biol. 1979;49:127–186. [PubMed] [Google Scholar]
  • Lester G. Regulation of tryptophan biosynthetic enzymes in Neurospora crassa. J Bacteriol. 1971 Jul;107(1):193–202.[PMC free article] [PubMed] [Google Scholar]
  • SUYAMA Y, LACY AM, BONNER DM. A GENETIC MAP OF THE TD LOCUS OF NEUROSPORA CRASSA. Genetics. 1964 Jan;49:135–144.[PMC free article] [PubMed] [Google Scholar]
  • KAPLAN S, SUYAMA Y, BONNER DM. FINE STRUCTURE ANALYSIS AT THE TD LOCUS OF NEUROSPORA CRASSA. Genetics. 1964 Jan;49:145–158.[PMC free article] [PubMed] [Google Scholar]
  • Metzenberg RL, Stevens JN, Selker EU, Morzycka-Wroblewska E. Identification and chromosomal distribution of 5S rRNA genes in Neurospora crassa. Proc Natl Acad Sci U S A. 1985 Apr;82(7):2067–2071.[PMC free article] [PubMed] [Google Scholar]
  • Matchett WH, DeMoss JA. The subunit structure of tryptophan synthase from Neurospora crassa. J Biol Chem. 1975 Apr 25;250(8):2941–2946. [PubMed] [Google Scholar]
Department of Biological Sciences, Stanford University, Stanford, CA 94305
Present address: Codon, 430 Valley Drive, Brisbane, CA 94005.
To whom reprint requests should be addressed.
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Abstract
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.
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