Low-frequency chimeric yeast artificial chromosome libraries from flow-sorted human chromosomes 16 and 21.
Construction of chromosome-specific yeast artificial chromosome (YAC) libraries from sorted chromosomes was undertaken (i) to eliminate drawbacks associated with first-generation total genomic YAC libraries, such as the high frequency of chimeric YACs, and (ii) to provide an alternative method for generating chromosome-specific YAC libraries in addition to isolating such collections from a total genomic library. Chromosome-specific YAC libraries highly enriched for human chromosomes 16 and 21 were constructed. By maximizing the percentage of fragments with two ligatable ends and performing yeast transformations with less than saturating amounts of DNA in the presence of carrier DNA, YAC libraries with a low percentage of chimeric clones were obtained. The smaller number of YAC clones in these chromosome-specific libraries reduces the effort involved in PCR-based screening and allows hybridization methods to be a manageable screening approach.
McCormick
M K
MK
Life Sciences Division, University of California, M880 Los Alamos National Laboratory, NM 87544.
Campbell
E
E
Deaven
L
L
Moyzis
R
R
D002458:Cell Fractionation; D002874:Chromosome Mapping; D015825:Chromosomes, Fungal; D002877:Chromosomes, Human; D002885:Chromosomes, Human, Pair 16; D002891:Chromosomes, Human, Pair 21; D003001:Cloning, Molecular / Q000379:methods*; D004247:DNA / Q000302:isolation & purification; D005434:Flow Cytometry / Q000379:methods; D015723:Gene Library; D006801:Humans; D006822:Hybrid Cells; D017403:In Situ Hybridization; D007621:Karyotyping; D016133:Polymerase Chain Reaction; D014170:Transformation, Genetic
D004247:DNA