Conditional genomic rearrangement by designed meiotic recombination using VDE (PI-SceI) in yeast.
Meiotic recombination plays critical roles in the acquisition of genetic diversity and has been utilized for conventional breeding of livestock and crops. The frequency of meiotic recombination is normally low, and is extremely low in regions called "recombination cold domains". Here, we describe a new and highly efficient method to modulate yeast meiotic gene rearrangements using VDE (PI-SceI), an intein-encoded endonuclease that causes an efficient unidirectional meiotic gene conversion at its recognition sequence (VRS). We designed universal targeting vectors, by use of which the strain that inserts the VRS at a desired site is acquired. Meiotic induction of the strains provided unidirectional gene conversions and frequent genetic rearrangements of flanking genes with little impact on cell viability. This system thus opens the way for the designed modulation of meiotic gene rearrangements, regardless of recombinational activity of chromosomal domains. Finally, the VDE-VRS system enabled us to conduct meiosis-specific conditional knockout of genes where VDE-initiated gene conversion disrupts the target gene during meiosis, serving as a novel approach to examine the functions of genes during germination of resultant spores.
Fukuda
Tomoyuki
T
Genetic System Regulation Laboratory, RIKEN, 2-1 Hirosawa, Wako, Saitama, 351-0198, Japan. tfukuda@riken.jp
Ohya
Yoshikazu
Y
Ohta
Kunihiro
K
D002470:Cell Survival / Q000235:genetics; D053903:DNA Breaks, Double-Stranded; D032621:DNA Shuffling / Q000379:methods*; D018390:Gene Targeting / Q000379:methods; D016681:Genome, Fungal; D042822:Genomic Instability / Q000235:genetics; D008540:Meiosis / Q000235:genetics; D008954:Models, Biological; D016254:Mutagenesis, Insertional; D030781:Organisms, Genetically Modified; D006180:Proton-Translocating ATPases / Q000502:physiology*; D011995:Recombination, Genetic; D029701:Saccharomyces cerevisiae Proteins / Q000502:physiology*; D013172:Spores, Fungal / Q000166:cytology / Q000235:genetics; D015003:Yeasts / Q000235:genetics*
D029701:Saccharomyces cerevisiae Proteins; D006180:Proton-Translocating ATPases; C081687:VMA1 protein, S cerevisiae