A set of transgenic insertion stocks derived by TE mobilization using the Tni\piggyBac-based construct PBac{RB}. The PBac{RB} construct carries the w+mC mini-white marker, a single long (199-bp) Scer\FRT site, and a "splice trap," a splice acceptor site derived from a D. melanogaster gene. The FRT site allows Scer\FLP-mediated recombination between other FRT-containing elements, and thus can be used to generate molecularly defined deletions.
PBac{RB} insertion lines from Exelixis were remapped and assessed for inclusion in the Gene Disruption Project collection; flanking sequence data were submitted to GenBank.
The PBac{RB} element was mobilized using P{Tub-PBac\T} as the source of Tni\piggyBac transposase. All stocks used for creation and characterization of new insertions were in an isogenic w[-] background. Both dysgenic males and females were used; from the dysgenic crosses a single new insertion was retained per vial, to avoid premeiotic clusters. Insertions were mapped to a chromosome by standard genetic methods, examined for homozygous viability and used for recovery of flanking genomic sequence.
Markers in stocks (as originally submitted to the BDSC):w1118.
Flanking genomic sequences from both 5′ and 3′ piggyBac ends were obtained by sequencing of inverse PCR products, and were mapped by alignment to the genomic sequence. For the combined Exelixis lines, 89% could be uniquely mapped to the genome.
Reanalysis of the genomic mapping of most of the combined Exelixis collections (insertions of P{XP}, PBac{PB}, PBac{RB}, and PBac{WH}) was undertaken. After multiple assessments, including sequencing and alignment of inverse PCR products, 16,073 insertions were mapped to unique sites. Of these, 1859 lines were added to the Gene Disruption Project collection.
