title: Analysis of a clonal selection event during transposon-mediated nested-deletion formation in rare BAC and PAC clones.
abstract: Nested deletions from one end of the genomic DNA in bacterial artificial chromosomes (BACs) and P1 artificial chromosomes (PACs) are readily generated by inserting a loxP site-containing Tn10 minitransposon into the recombinant clone and transducing with P1 phage. Although the size of clones in the deletion series is largely random, in about 5% of BACs and PACs the distribution appears skewed to a certain length, and in rare cases (<1%) is definitely skewed to a particular size. Here we investigate this relatively rare phenomenon and validate that sequence-specific transposon insertions are not the cause of such skewed nested-deletion libraries. Instead, a detailed analysis of our experiments with a BAC clone demonstrating this unusual feature indicates that deletions of a certain size arise from clonal expansion of a transposon insertion as a result of transient derepression of the transposase gene prior to IPTG induction. Transposition itself shows no bias to any particular region of insert DNA in the clone. We suggest a simple modification to the procedure for generating nested-deletions that allows all BACs and PACs to produce nested-deletions of random size. These findings should provide additional insight into the causes of site selectivity in genomic clones with other inducible transposon systems.
authors.affiliation: Biomedical/Biotechnology Research Institute, North Carolina Central University, 1801 Fayetteville Street, Durham, North Carolina 27707, USA. pchatterjee@wpo.nccu.edu
authors.forename: P K
authors.initials: PK
authors.lastname: Chatterjee
authors.forename: L P
authors.initials: LP
authors.lastname: Briley
mesh_terms: D022201:Chromosomes, Artificial; D022202:Chromosomes, Artificial, Bacterial; D004251:DNA Transposable Elements; D007544:Isopropyl Thiogalactoside / Q000737:chemistry; D017384:Sequence Deletion
chemical_list: D004251:DNA Transposable Elements; D007544:Isopropyl Thiogalactoside