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. 2016 May 2;113(20):5622–5627. doi: 10.1073/pnas.1600108113

Fig. S1.

Fig. S1.

CRISPR design and genotyping for knockout and EmGFP tagging of the TPCN2 gene, coding for the TPC2 protein. (A) Schematic of the TPC2 protein topology. Red stars indicate the two amino acid changes caused by single nucleotide polymorphisms strongly associated with pigmentation variations in a genome-wide association study (13). (B) Schematic of the CRISPR/Cas9 design. (C, Left) Genotyping of TPC2-KO CRISPR clones. PCR using the indicated primers (OUT-Fwd-N and OUT-Rev-N) was carried out for mock transfected MNT-1 cells and MNT-1 homozygous clones (TPC2-KO1 and TPC2-KO2), obtained by transfection with the indicated sgRNAs (sgRNA1+2 and sgRNA1+3, respectively). (Right) Genotyping of a heterozygous TPC2-EmGFP CRISPR clone. PCR using the indicated primers (OUT-Fwd-C and OUT-Rev-C primers or EmGFP primers) was carried out for mock-transfected MNT-1 cells and MNT-1 cells transfected with the indicated sgRNAs (sgRNA4+5) and donor plasmid.