Figure 1

Sstr3::GFP ROSA26 targeted alleles. (A) Schematic of the Sstr3::GFP conditional (Cilia^GFP-OFF) and expressing (Cilia^GFP-ON) alleles. Before Cre mediated deletion (Cilia^GFP-OFF), the expression of Sstr3::GFP is precluded by the PGK promoted Neo cassette flanked by LoxP sites (blue triangles). After Cre mediated recombination (Cilia^GFP-ON) at the LoxP sites the Neo cassette is deleted allowing the expression of Sstr3::GFP. Splice acceptor (SA), blue triangles represent LoxP sites, PKG-Neo: Phosphoglycerate kinase promoter driving a neomycin resistance gene followed by three polyA tail signals (3XpA, black). The remaining attB sites after Gateway recombination are represented by maroon boxes flanking the cDNA of Sstr3::GFP. (B) To confirm expression and ciliary localization, embryonic stem (ES) cells with correctly targeted Sstr3::GFP were differentiated and transduced with a Cre expressing lentivirus to induce expression of Sstr3::GFP. (C) A four primer combination was used for genotyping by PCR to amplify the wild type (226 bp), Cilia^GFP-OFF (317 bp) and Cilia^GFP-ON (423 bp) alleles. Lane 1:100 bp ladder, lane 2: Cilia^GFP-OFF/Cilia^GFP-OFF, lane 3: Cilia^GFP-OFF/WT, and lane 4: Cilia^GFP-ON/WT. Primer binding sites represented in (A) as small black arrows, sequences in Methods.