Drosophila promoters

These vectors have the Ubiquitin promoter with optional N-terminal epitope tags following the initial ATG. We have used this promoter with fusions to cDNAs of Mad to rescue maternal and zygotic phenotypes (see Sekelsky et al. 1995) and mei-9 to rescue phenotypes in somatic cells and female meiosis (see Yildiz et al. 2004). The pBU series are in pBlueScript KS+; the pWU vectors are in our pP{70w} P element transformation vector.

pBUF

FLAG epitope (or no tag)

Map

Sequence

GCK2 file

pBUHA

HA epitope

Map

Sequence

GCK2 file

pBUSH

Six histidine tag

Map

Sequence

GCK2 file

pWUM6

Six Myc epitopes

Map

Sequence

GCK2 file

pWUM6b

Six Myc epitopes

Map

Sequence

GCK2 file

Drosophila gene targeting vectors

See also our Guide to Gene Targeting (which is way out of date) and a list of References reporting successful targeting with these vectors.
 

The pP{Target} series vectors are for use in the ends-in targeted gene replacement method of Rong and Golic. Each has a polylinker, white marker gene, and I-CreI site, flanked by FRT sites and P element ends. These vectors all have minimal FRT sites - only 34 bp. In our hands, these minimal FRTs are as efficient as the longer FRTs shown to work well by Kent Golic and colleagues.

pP{Target}

mini-white from CaSpeR

Map

Sequence

GCK2 file

pP{TargetB}

Smaller version of above

Map

Sequence

GCK2 file

pP{Target70}

hsp70::white cDNA

Map

Sequence

GCK2 file

pP{Target70i}

hsp70::white cDNA, with intron

Map

Sequence

GCK2 file

Rong and Golic used "ends-in" (insertional) targeting, and Gong and Golic showed that "ends-out" (gene replacement) is also feasible. Our vector for this method is pP{EndsOut}. A genomic region is cloned into the polylinker, and the gene of interest is disrupted or replaced with the hsp70::white cDNA, which also serves as a marker for transformation. To reduce the size of this construct, we put the FRT sites adjacent to the I-sites. Sometimes, I-SceI will cut and imprecise repair will result in loss of the I-SceI cut sequence. A small subset of these cases might involve deletion into the FRT site. If this happens, the FRT will not be usable, and you may get a "false positive" that represents and unexcisable donor. We estimate that this typically occurs on the order of once in 10⁴-10⁵ progeny, so it should not be a majror source of false positives, except in those targeting experiments that have low efficiency.

pP{EndsOut2}

For ends-out gene replacement

Map

Sequence

GCK2 file

pBS-70w

hsp70-white cDNA

Map

Sequence

GCK2 file

pP{whiteOut2}

For ends-out using white for transformation,
but not for selecting targeted events

Map

Sequence

GCK2 file