Genomic reconstruction by serial mitotic recombination of yeast artificial chromosomes.
DNA cloned in yeast artificial chromosomes (YACs) is a valuable resource for functional experiments in cell culture as well as whole animal systems. Where the size or chimerism of a YAC clone are limiting factors it may be desirable to generate recombinant YAC clones. One such approach is based on mitotic recombination, and we describe the development of a methodology that allows multiple recombination cycles for serial reconstruction of overlapping YACs. This approach employs retrofitting with standard plasmid vectors, transfer of YACs to a common haploid host by Kar1 mating, and selection for recombination with 5-fluoro-orotic acid.
Markie
David
D
Pathology Department, Dunedin School of Medicine, New Zealand.
Jones
Emma
E
Ragoussis
Jiannis
J
D018244:Chromosomes, Artificial, Yeast; D005819:Genetic Markers; D005822:Genetic Vectors; D016678:Genome; D006238:Haploidy; D008938:Mitosis / Q000235:genetics*; D009963:Orotic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D010957:Plasmids; D011995:Recombination, Genetic; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D012641:Selection, Genetic
D005819:Genetic Markers; D009963:Orotic Acid