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. 2021 Oct 21;16(11):2768–2783. doi: 10.1016/j.stemcr.2021.09.019

Figure 1.

Figure 1

MPN-related iPSC clones exhibit pluripotency state and harbor CALR mutations

(A) Pluripotency assessment of indicated iPSC clones by immunofluorescence. Merge represents overlay of DAPI (blue), OCT3/4 (green), and TRA-1-60 (red). Scale bars, 100 μm.

(B) CALR mRNA expression of indicated iPSC clones was assessed by CALR mutant allele-specific qRT-PCR. Gene expression is depicted as percentage of GAPDH. Mean value ± SD of representative clones with indicated CALR genotypes, n = 2 independent experiments.

(C) Mutant CALR expression (red) was confirmed by immunofluorescence staining with a CALR mutant specific antibody. Unmutated CALR iPSCs served as negative control. Nuclei were stained with DAPI (blue). Scale bars, 100 μm.

(D) Representative western blot analysis of CALR WT and CALR mutant (Mut) protein in iPSC clones after CRISPR repair of hom CALRins5 and hom CALRdel52 mutations. CALR WT protein was assessed on the same membrane as CALR mutant without stripping of the CALR Mut antibody explaining residual CALR mutant bands. GAPDH was used as loading control.