Reprogramming JPCs to iPSCs via VEE-OKSM-GFP RNA

JPCs were reprogrammed to iPSCs as previously published [13 (link)]. Briefly, JPCs were incubated in hPL5 medium containing 0.2 µg/mL recombinant B18R protein (eBioscience, San Diego, CA, USA) prior to transfection with a self-replicating VEE-OKSM-GFP RNA. From day 1–5 transfected cells were incubated with hPL5 + 25% B18R conditioned medium (BcM) + 1 µg/mL puromycin (Invivogen, Toulouse, France) to select transfected cells. Total of 250 µM histone deacetylase inhibitor sodium butyrate (NaB, Selleck Chemicals LLC, Houston, TX, USA) was added to the medium to enhance reprogramming efficiency. At day 7, the medium was changed to E8 medium (Essential 8, Thermo Fisher Scientific Inc., Waltham, USA) + 25% BcM. Single iPSC colonies were picked and transferred into VTN-coated wells containing E8 medium + 10 µM Y27632 ROCK inhibitor (Selleck Chemicals LLC, Houston, TX, USA) and maintained in E8 medium with daily medium changes. Passaging was performed every 4–6 days.