Figure 1

Generating panx1a^−/− fish using TALENs – (a) The zebrafish panx1a gene structure with six coding exons. The position of the left and right TALENs sequence with the spacer sequence and AfeI restriction site in blue and red is highlighted. (b) Larval survival rates (in %) one day after (1 dpf) microinjection. A dose of 30 pg TALENs pair resulted in more than 50% survival rate and was selected for the experiments generating the panx1a^−/− fish line. (c) The RFLP-assay shows the loss of the AfeI recognition sequence (indicated by uncut) in four out of ten F0 larvae tested. (d) A sequencing alignment is demonstrating small deletions causing frameshift mutations found in three different F0 larvae. (e) The predicted sequence features of the mutated panx1a were visualized using the Protter open-source tool (wlab.ethz.ch/protter). A 4 bp deletion in Panx1a exon 4 resulted in a frameshift causing a premature stop codon at amino acid 195. (f) Localization of the truncated Panx1a protein – Confocal images of transiently transfected proteins Panx1a^wt-EGFP (left panel) and Panx1a^Δ4-EGFP (right panel) in Neuro2a cells. Nuclei were stained with DAPI. Scale bar: 10 μm.