Fig. 3

Rescue of the jaguar/obelix Phenotype by BAC Injection and Expression of Kir7.1 mRNA. (A) Line diagram depicting the genomic locations of the putative Kir7.1 gene and BAC clones used for the rescue experiments. We used two BAC clones (98K222and 126F9, represented by gray lines) for microinjection. Numbers below the thick black line indicate the positions of the BACs and Kir7.1 on LG15. Open boxes with vertical lines represent the positions and intron-exon structures of putative genes predicted from the ENSEMBL transcript. (B) BAC rescue of pigment pattern in zebrafish mutants. Fertilized eggs from homozygous mutant fish (jag^b230 and obe^td15) were used in the phenotype rescue experiment. For the injected fish that survived to adulthood, representative pigment patterns of whole body (top), trunk (middle), and anal fin (bottom) are shown. The left panels depict patterns resulting from phenotype rescue using BAC clone 98K222. The middle panels depict patterns resulting from phenotype rescue using BAC clone 126F9. The right panels depict patterns of noninjected fish. All mutant fish (jag^b230 and obe^td15) rescued by BAC injection had a partial stripe patterns. (C) PCR analysis to confirm BAC integration in the rescued fin. Agarose gel analysis of PCR products derived from a BAC-specific sequence in nonrescued fins (lanes 2 and 3) and rescued fins (lanes 4 and 5). DNA fragments derived from the BAC clones were detected in all rescued fish (n = 6), but no PCR fragment was obtained from DNA of nonrescued fish (n = 6). Molecular mass markers are indicated (m). (D) Expression of Kir7.1 mRNA as detected by single-cell RT-PCR. Agarose gel analysis of RT-PCR products derived from individual melanophore (M), xanthophore (X), or fin dermal cells (F). Molecular mass markers are indicated (m).