Generation of Large Insert YAC Libraries -- YAC Library Storage and Transport -- YAC Library Screening I -- YAC Library Screening II -- Cloning of Human Telomeres in Saccharomyces cerevisiae -- Purification of YAC-Containing Total Yeast DNA -- Restriction Analysis of YACs -- RARE-Cleavage Analysis of YACs -- YAC Localization by Fluorescence In Situ Hybridization -- Alu-PCR Fingerprinting of YACs -- Detection of Chimerism in YAC Clones -- Amplification with Arbitrary Primers -- End Rescue from YACs Using the Vectorette -- Isolation of YAC Ends by Plasmid Rescue -- End-Rescue of YAC Clone Inserts by Inverse PCR -- Covering YAC-Cloned DNA with Phages and Cosmids -- Fragmentation and Integrative Modification of YACs -- Targeting Mutations to YACs by Homologous Recombination -- Reconstruction of Large Genomic Segments of DNA by Meiotic Recombination Between YACs -- Genomic Reconstruction by Mitotic Recombination of YACs -- Amplification of the Copy Number of YACs -- Transfer of YAC Clones to New Yeast Hosts -- Use of ACEDB as a Database for YAC Library Data Management -- YAC Transfer into Mammalian Cells by Cell Fusion -- YAC Transfer by Microinjection -- Transfection of Mammalian Cells via Lipofection -- The Isolation of cDNAs by Hybridization of YACs to cDNA Libraries -- cDNA Selection with YACs -- Markers, Selection, and Media in YAC Cloning.

Yeast artificial chromosomes (YACs) have their origins in the molecular genetic analysis of the yeast Saccharomyces cerevisiae. The construction of self-maintaining genetic elements from isolated frag­ ments of the yeast genome defined DNA sequences necessary for chro­ mosome function has provided telomeres, centromeres, and autonomous replicating sequences. In 1987 a reversal of the strategy put these short functional DNA sequences to work in cloning vectors, producing "yeast" chromosomes largely composed of foreign DNA. Initially the insert size of clones averaged several hundred kilobasepairs, a remarkable achieve­ ment. Rapid progress with cloning technology has since enabled the construction of YAC libraries with average insert sizes of around 1 Mb, with many clones exceeding that size, and YACs remain the largest capacity microbiological cloning system available. They effectively bridge the size gap between bacterial cloning (plasmids, cosmids, PI, and bacterial artificial chromosomes) and what could be considered mammalian cloning systems (somatic cell hybrids and irradiati- fusion gene transfer hybrids). YACs also brought with them a conceptual revolution in the man­ agement of clone libraries. The large carrying capacity of YACs, with subsequent reduction in the total number required, meant that it was conceivable to store clones individually rather than as pools that require constant re-plating. Each clone in the library has a unique address and, with successive screenings, information accumulates about individual clones.