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Fig. 7 Standard RT-PCR for testing splice variants, exon skipping, and expression of pdzk1 in wild-type fish and pdzk1-KO mutants. (a) Schematics of the two primer pairs for pdzk1 mRNA and their products. Pdzk1 mRNA is expressed from the nine exons of pdzk1 genomic DNA. In mutants, the insertion (highlighted in gray on the pdzk1 mRNA sequence and primer products) should also be expressed within the exon 2 region (green) of the mRNA. Primer pair 1 (P1) was designed to amplify the mRNA region across exons 1 to 6 containing the insertion, producing 980-bp products for wild-type fish and longer (1008-bp) products for mutants due to the insertion. Primer pair 2 (P2) was designed to amplify the mutated pdzk1 mRNA sequence, which starts within the insertion in exon 2 and ends in exon 6. P2 produces 901-bp products only for mutants, with no product for wild-type controls. Orange ends of products represent primers. (b) As expected, standard RT-PCR using P1 produced slightly longer products for mutants than for wild-type fish. (c) RT-PCR using P2 produced ∼900-bp products for mutants but no amplified sequence for wild-type fish.