Figure 1

Generation and basic characterization of alpl^-/- zebrafish line. (a) Schematic representation of vitamin B₆ metabolism. (b) Sanger sequencing results showing a 10 bp deletion at the CRISPR site in exon 5 of the alpl gene (c.623_632del, p.Gln180fs), which is predicted to result in a truncated protein. (c) Agarose electrophoresis of genotyping PCR products for the CRISPR site in exon 5 of the alpl gene in wild type (WT), alp^+/-, alpl^-/- 5 dpf old zebrafish embryos. (d) Relative alpl mRNA expression in WT, alpl^+/-, and alpl^-/- 5 dpf old zebrafish embryos. Data are means from n = 3 pools (10 embryos per pool) per genotype measured in duplicate ± SD. **** p < 0.0001. (e) Total alkaline phosphatase activity in whole-embryo extracts of WT, alpl^+/-, and alpl^-/- 5 dpf old zebrafish. RLU, relative light unit. Data are means from n = 3 pools (10 embryos per pool) per genotype ± SD. **** p < 0.0001 and *** p < 0.001. (f) Alpl protein expression in total WT and alpl^-/- 5 dpf zebrafish embryo extracts showing lack of alpl protein in alpl^-/- embryos (predicted molecular weight of alpl protein (NP_957301.2) is 62.5 kDa). Data are from pools of n = 32 embryos per genotype. Gapdh protein expression was used as the loading control.