Hesc qualified geltrex basement membrane matrix

UABi001-A and UABi002-A were submitted to Cell Line Genetics (Madison, WI) for G-band karyotyping, which analyzes twenty G-banded metaphase cells of each cell line. Teratoma formation assay was conducted as described (Liu et al., 2015 ). Briefly, 1 × 10⁶–5 × 10⁶ iPSCs were suspended into 100 μl of DMEM/F12 medium containing 30% hESC-qualified GelTrex Basement Membrane Matrix (Thermo Fisher) and injected subcutaneously into six week-old NOD SCID gamma (NSG) mice (005557, The Jackson Laboratory). Tumors were harvested, fixed with 10% formalin, and submitted to the Comparative Pathology Laboratory at University of Alabama at Birmingham (UAB). All animal experiments were controlled under the guidelines from UAB and NIH.

STEMCCA lentivirus, which express all reprogramming factors from a polycistronic transcript [18 (link)], was prepared as described [19 (link)]. Primary LGG cells were transduced with the STEMCCA lentivirus at an MOI of 3 as described [20 (link)]. Three days after transduction, the LGG cells were reseeded onto cell culture plates-coated with hESC-qualified Geltrex basement membrane matrix (Thermo Fisher) at a density of 1−3×10⁵ cells/cm² in LGG medium. The cells were then cultured in E7 medium (E8 medium without TGF-β) from the following day for two weeks and in E8 medium for one to two weeks as described [20 (link)–22 (link)]. Independent clones of iPSCs were manually picked into Geltrex-coated 12-well tissue culture plates and maintained in E8 medium as described [23 (link),24 (link)].