RRC ID 12230
Author Wada M, Abe K, Okumura K, Taguchi H, Kohno K, Imamoto F, Schlessinger D, Kuwano M.
Title Chimeric YACs were generated at unreduced rates in conditions that suppress coligation.
Journal Nucleic Acids Res
Abstract Chimerism is a major limitation of current YAC libraries. A method based on partially filled-in ends of restriction fragments was designed to avoid coligation as a possible source of chimeras. Model experiments using plasmid DNA as an insert showed that coligation was clearly avoided by this method. Pilot collections of YACs with an average insert size of 650kb were then constructed with and without the partial fill-in treatment. Starting from a mixture of a equal amounts of human and mouse DNA, none of 108 clones was positive by hybridization with both Alu and B2 probes, again suggesting that coligation was effectively blocked. However, 4 out of 10 clones still hybridized to 2 or more locations by FISH on chromosomes in human metaphase spreads, level similar to that in the clones made without the partial fill-in step. These results strongly suggest that chimeric clones generally arise by a mechanism independent of coligation, presumptively based on recombination.
Volume 22(9)
Pages 1651-4
Published 1994-5-11
DOI 10.1093/nar/22.9.1651
PMID 8202367
PMC PMC308044
MeSH Animals Cell Line Chromosomes, Artificial, Yeast* DNA, Recombinant Gene Library Humans Kinetics Mice Plasmids Sepharose
IF 11.502
Times Cited 14
DNA material pBLUR8 (RDB01029)