Transgene-Free Production of Pluripotent Stem Cells Using piggyBac Transposons

Reprogramming of somatic cells to induced pluripotent stem cells (iPSCs) allows the derivation of �personalized stem cells. Transposon transgenesis is a novel and viable alternative to viral transduction methods for the delivery of reprogramming factors (Oct4 , Sox2 , Klf4 , c-Myc ) to somatic cells. Since transposons can be introduced as naked DNA using common plasmid transfection protocols, they provide a safer alternative to viral methods. piggyBac transposons are host-factor independent and integrate stably into the target genome, yet benefit from the unique characteristic of seamless removal mediated by transient expression of piggyBac transposase. Thus, piggyBac transposition provides an effective means to generate human, transgene-free iPSCs. The protocol describes the production of iPSCs from human embryonic fibroblasts, delivering reprogramming factors via plasmid transfection and piggyBac transposition.