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Nucleic Acids Res

Title:		Stabilization of yeast artificial chromosome clones in a rad54-3 recombination-deficient host strain
Author(s):		Le Y; Dobson MJ;
Address:		"Department of Biochemistry, Faculty of Medicine, Sir Charles Tupper Medical Building, Dalhousie University, Halifax, Nova Scotia B3H 4H7, Canada"
Journal Title:		Nucleic Acids Res
Year:		1997
Volume:		25
Issue:		6
Page Number:		1248 - 1253
DOI:		10.1093/nar/25.6.1248
ISSN/ISBN:		0305-1048 (Print) 1362-4962 (Electronic) 0305-1048 (Linking)
Abstract:		"The cloning and propagation of large fragments of DNA on yeast artificial chromosomes (YACs) has become a routine and valuable technique in genome analysis. Unfortunately, many YAC clones have been found to undergo rearrangements or deletions during the cloning process. The frequency of transformation-associated alterations and mitotic instability can be reduced in a homologous recombination-deficient yeast host strain such as a rad52 mutant. RAD52 is one member of an epistatic group of genes required for the recombinational repair of double-strand breaks in DNA. rad52 mutants grow more slowly and transform less efficiently than RAD + strains and are therefore not ideal hosts for YAC library construction. We have investigated the ability of both null and temperature-sensitive alleles of RAD54 , another member of the RAD52 epistasis group, to prevent rearrangements of human YAC clones containing tandemly repeated DNA sequences. Our results show that the temperature-sensitive rad54-3 allele blocks mitotic recombination between tandemly repeated DYZ3 satellite sequences and significantly stabilizes a human DYZ5 satellite-containing YAC clone. Yeast carrying the rad54-3 mutation can undergo meiosis, have growth and transformation rates comparable with RAD + strains, and therefore represent improved YAC cloning hosts"
Keywords:		"*Chromosomes, Artificial, Yeast DNA Helicases DNA Repair Enzymes DNA, Satellite/genetics Fungal Proteins/*genetics/metabolism Gene Library Genotype Humans Mating Factor Meiosis Mitosis Mutagenesis Peptides/genetics *Recombination, Genetic *Repetitive Sequ;"
Notes:		"MedlineLe, Y Dobson, M J eng Research Support, Non-U.S. Gov't England 1997/03/15 Nucleic Acids Res. 1997 Mar 15; 25(6):1248-53. doi: 10.1093/nar/25.6.1248"