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. Author manuscript; available in PMC: 2019 Sep 10.
Published in final edited form as: Stem Cell Res. 2019 Jul 16;39:101504. doi: 10.1016/j.scr.2019.101504

Figure 1. SCA3-hESC line UM134–1 is pluripotent, possesses a normal karyotype, and expresses pathogenic polyglutamine-expanded mutant ATXN3.

Figure 1.

(A) Immunocytochemical analysis with DAPI co-stain of SCA3-hESC revealed expression of pluripotency markers OCT3/4, SOX2, and NANOG. Scale bar = 200 μm. (B) Undifferentiated WT-hESC (blue) and SCA3-hESC (red) expressed pluripotency markers LIN28, OCT4, SOX2 and NANOG as assessed by qRT-PCR analysis. Data are represented as mean of three replicates ± SEM. (C) SCA3-hESC were differentiated into embryoid bodies for 21 days in culture. Differentiated SCA3-embryoid bodies expressed lineage markers of endodermal [α-fetoprotein (AFP)], mesodermal [Brachyury], and ectodermal tissue [neuron-specific class III beta-tubulin (TUJ-1)]. Electrophoresis demonstrated anticipated amplicon size for each lineage marker PCR primer set. (D) G-banded karyotype analysis of passage 6 undifferentiated SCA3-hESC showed a normal 46,XY karyotype. (E) Representative anti-polyQ expansion and (F) anti-ATXN3 Western blot of undifferentiated WT- and SCA3-hESC revealed heterozygous expression of polyQ-expanded ATXN3 protein in SCA3-hESC within the pathogenic range for SCA3. (G) Anticipated ATXN3 polyQ repeat lengths in WT- and SCA3-hESC lines as determined by gene fragmentation analysis. Mutant polyQ-expanded repeat length is highlighted in red. (mutATXN3 = mutant ATXN3; wtATXN3 = wild type ATXN3).