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Episomal transgene expression in pluripotent stem cells.

Methods in molecular biology (Clifton, N.J.) (2011-08-09)
Michele M P Lufino, Anna R Popplestone, Sally A Cowley, Pauline A H Edser, William S James, Richard Wade-Martins
ABSTRACT

Herpes simplex type 1 (HSV-1) amplicon vectors possess a number of features that make them excellent vectors for the delivery of transgenes into stem cells. HSV-1 amplicon vectors are capable of efficiently transducing both dividing and nondividing cells and since the virus is quite large, 152 kb, it is of sufficient size to allow for incorporation of entire genomic DNA loci with native promoters. HSV-1 amplicon vectors can also be used to incorporate and deliver to cells a variety of sequences that allow extrachromosomal retention. These elements offer advantages over integrating vectors as they avoid transgene silencing and insertional mutagenesis. The construction of amplicon vectors carrying extrachromosomal retention elements, their packaging into HSV-1 viral particles, and the use of HSV-1 amplicons for stem cell transduction will be described.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Ethylenediaminetetraacetic acid disodium salt dihydrate, suitable for electrophoresis, Molecular Biology, 99.0-101.0% (titration)
Sigma-Aldrich
Y-27632-CAS 331752-47-7-Calbiochem, Y-27632A, CAS 331752-47-7, is a cell-permeable, reversible, inhibitor of Rho kinases (Ki = 140 nM for p160ROCK). Enhances survival & cloning efficiency of ESC without affecting their pluripotency.