Extended Data Fig. 4: Generation of SLC30A8-p.Lys34Serfs50* and SLC30A8-p.Arg138* hiPSC lines.
From: Loss of ZnT8 function protects against diabetes by enhanced insulin secretion
a, CRISPR-Cas9 strategy to generate SLC30A8-p.Lys34Serfs50* (Exon 2) and SLC30A8-p.Arg138* (Exon 3) hiPSC lines. Orange font highlights the nucleotide changes: c.101-107del; p.Lys34Serfs50* and c.412C>T; p.Arg138*. The gRNA (blue font) and PAM sequences (red font) are indicated on the partial genomic sequence of SLC30A8. b-c, FACS data from undifferentiated b, SLC30A8-p.Arg138* and c, SLC30A8-p.Lys34Serfs50* iPSCs and relevant isotype controls using antibodies against: OCT3/4, SSEA, SOX2, and NANOG. d, Expression of INSULIN in hiPSC-derived beta-like cells. Black bars represent p.Arg138Arg control cells, red bars represent p.Arg138*, and yellow bars represent p.Lys34Serfs50*. (n=6-8 wells from three differentiations) e-g, RNAscope analysis of the number of e, INSULIN- and f, SLC30A8- transcript positive cells in hiPSC-derived beta-like cells. 7-21 image fields were quantified and presented as % of DAPI+ cells. Representative images used for quantification shown in g (scale bar = 50 µm). Data are presented as Mean±SEM. Statistical analysis was performed using the one-way ANOVA and Tukey’s multiple comparison test (n = 5-9 wells from three differentiations, ****p<0.0001).
