Designing and testing chimeric zinc finger transposases.
Transposons have been effectively utilized as non-viral gene delivery systems that are capable of promoting stable transgene expression in mammalian and human cells. Two specific transposon systems, Sleeping Beauty (SB) and piggyBac (PB), have been successfully modified by the addition of a zinc finger (ZF) DNA-binding domain as a strategy for directing transposon integration near ZF target sites in a host cell genome. Site-directed integration could improve transposon-mediated gene transfer by limiting positional effects on transgene integration, limiting genotoxicity, and improving the overall safety in gene therapy applications. In this chapter, we describe methodology for creating and characterizing functional chimeric ZF transposases and experimental approaches for testing their transpositional activity in human cells.
Wilson
Matthew H
MH
Department of Medicine, Michael E. DeBakey VA Medical Center, Baylor College of Medicine, Houston, TX, USA.
George
Alfred L
AL
D015153:Blotting, Western; D002460:Cell Line; D004251:DNA Transposable Elements / Q000235:genetics; D006801:Humans; D008957:Models, Genetic; D008962:Models, Theoretical; D016297:Mutagenesis, Site-Directed; D019895:Transposases / Q000235:genetics / Q000378:metabolism*; D016335:Zinc Fingers / Q000235:genetics*
D004251:DNA Transposable Elements; D019895:Transposases