SBI offers a non-viral system utilising PhiC31 integrase for the reprogramming of somatic cells to a pluripotent state. Shown to be especially effective for generation of murine iPSCs, this highly efficient system is used widely in gene therapy applications, as the single copy integrated cassette includes a unique PinPoint site which can be used for targeted gene addition.
phiC31 Integrase iPSC Vectors
The pCOBLW phiC31 reprogramming vector features iPSC factors driven by a potent CAGs promoter with the factors separated by P2A elements. After co-transfection of the reprogramming vector with the phiC31 expression plasmid (catalog# FC200PA-1) and integration, monitor for successful transfection with GFP fluorescence& select for positive colonies using the built-in Neo marker. The reprogramming iPSC factor donor vector & the phiC31 expression plasmid maps are shown below.
phiC31 Integrase iPSC Lines - Example Data
Mouse induced pluripotent stem cells (iPSCs, Cat# SC211A-iPSC) were generated by nucleofecting genetically unmodified C57BL/6 mouse embryonic fibroblasts with the pCOBLW phiC31 integrase mouse reprogramming donor plasmid (Cat# FC305A-1) & the phiC31 integrase expression plasmid (Cat# FC200PA-1). Cells were derived using morphological selection criteria &GFP expression without drug selection. When cultured under standard mouse ES cell culture conditions, the morphology of the mouse iPSCs are identical to that of mouse ES cells. The cells also express the pluripotency markers SSEA-1, Nanog, Sox2 & Oct4 & demonstrate strong endogenous AP activity
The reprogramming donor plasmid encodes 4 murine transcription factors (Oct4, Sox2, Klf4, and c-Myc) shown to induce the reprogramming of somatic cells to a pluripotent state. Due to the specificity of recombination using the phiC31 integrase system, the iPSCs are expected to have, on average, one copy of the OSKM plasmid per cell, which reduces the risk of any insertional mutagenesis effects.
Material Available for Download
phiC31 Integrase iPSC Technology & Cell Lines User Manual
Note: product availability depends on country - see product detail page.
|Details||Cat number & supplier||Size||Price|
|PhiC31 Integrase Expression Plasmid FC200PA-1 · System Biosciences||
|Non-Viral PhiC31 Integrase Mouse iPSC Reprogramming System (includes FC200PA-1 and FC305A-1) FC300A-1 · System Biosciences||
|pCOBLW [CAG-Oct4-Sox2-Klf4-Myc-GFP-SV40-Neo] Mouse Reprogramming PhiC31 Donor Plasmid FC305A-1 · System Biosciences||