Pluripotent stem cells were derived from human fibroblasts using non-integrating episomal vectors. Cells express pluripotency markers, and acquired a normal karyotype 46 X,Y.
Human fibroblasts (106) were electroporated with pCXLE–hOCT3/4–shp53–F (or pCXLE-hOCT3/4), pCXLE–hSK and pCXLE–hUL, using a Nucleofector in combination with the NHDF nucleofector kit. After 6 days, fibroblasts were reseeded onto mitotically inactivated mouse embryonic fibroblasts (MEFs) and cultured in human embryonic stem cell (hESC) medium. The induced pluripotent stem cell (iPSC) colonies were allowed to grow before being mechanically picked and transferred onto MEFs. After successful expansion of the iPSC lines on MEFs, they were transferred to Matrigel and cultured in mTeSR medium.