Chapter title |
Zinc-finger nucleases-based genome engineering to generate isogenic human cell lines.
|
---|---|
Chapter number | 8 |
Book title |
Synthetic Gene Networks
|
Published in |
Methods in molecular biology, January 2012
|
DOI | 10.1007/978-1-61779-412-4_8 |
Pubmed ID | |
Book ISBNs |
978-1-61779-411-7, 978-1-61779-412-4
|
Authors |
Anne-Kathrin Dreyer, Toni Cathomen, Dreyer, Anne-Kathrin, Cathomen, Toni |
Abstract |
Customized zinc-finger nucleases (ZFNs) have developed into a promising technology to precisely alter mammalian genomes for biomedical research, biotechnology, or human gene therapy. In the context of synthetic biology, the targeted integration of a transgene or reporter cassette into a "neutral site" of the human genome, such as the AAVS1 locus, permits the generation of isogenic human cell lines with two major advantages over standard genetic manipulation techniques: minimal integration site-dependent effects on the transgene and, vice versa, no functional perturbation of the host-cell transcriptome. Here we describe in detail how ZFNs can be employed to target integration of a transgene cassette into the AAVS1 locus and how to characterize the targeted cells by PCR-based genotyping. |
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