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Zinc-finger nucleases (ZFNs) are versatile reagents that have redefined genome engineering. Realizing the full potential of this technology requires the development of safe and effective methods for delivering ZFNs into cells. We demonstrate the intrinsic cell-penetrating capabilities of the standard ZFN architecture and show that direct delivery of ZFNs as(More)
Transcription activator-like (TAL) effector nucleases (TALENs) have enabled the introduction of targeted genetic alterations into a broad range of cell lines and organisms. These customizable nucleases are comprised of programmable sequence-specific DNA-binding modules derived from TAL effector proteins fused to the non-specific FokI cleavage domain.(More)
Site-specific recombinases are tremendously valuable tools for basic research and genetic engineering. By promoting high-fidelity DNA modifications, site-specific recombination systems have empowered researchers with unprecedented control over diverse biological functions, enabling countless insights into cellular structure and function. The rigid target(More)
Zinc-finger recombinases (ZFRs) represent a potentially powerful class of tools for targeted genetic engineering. These chimeric enzymes are composed of an activated catalytic domain derived from the resolvase/invertase family of serine recombinases and a custom-designed zinc-finger DNA-binding domain. The use of ZFRs, however, has been restricted by(More)
The serine recombinases are a diverse family of modular enzymes that promote high-fidelity DNA rearrangements between specific target sites. Replacement of their native DNA-binding domains with custom-designed Cys₂-His₂ zinc-finger proteins results in the creation of engineered zinc-finger recombinases (ZFRs) capable of achieving targeted genetic(More)
Despite recent advances in genome engineering made possible by the emergence of site-specific endonucleases, there remains a need for tools capable of specifically delivering genetic payloads into the human genome. Hybrid recombinases based on activated catalytic domains derived from the resolvase/invertase family of serine recombinases fused to Cys2-His2(More)
Broadly neutralizing antibodies PG9 and PG16 effectively neutralize 70 to 80% of circulating HIV-1 isolates. In this study, the neutralization abilities of PG9 and PG16 were further enhanced by bioconjugation with aplaviroc, a small-molecule inhibitor of virus entry into host cells. A novel air-stable diazonium hexafluorophosphate reagent that allows for(More)
The construction of increasingly sophisticated synthetic biological circuits is dependent on the development of extensible tools capable of providing specific control of gene expression in eukaryotic cells. Here, we describe a new class of synthetic transcription factors that activate gene expression in response to extracellular chemical stimuli. These(More)
Targeted nucleases have provided researchers with the ability to manipulate virtually any genomic sequence, enabling the facile creation of isogenic cell lines and animal models for the study of human disease, and promoting exciting new possibilities for human gene therapy. Here we review three foundational technologies-clustered regularly interspaced short(More)
The development of new methods for delivering proteins into cells is a central challenge for advancing both basic research and therapeutic applications. We previously reported that zinc-finger nuclease proteins are intrinsically cell-permeable due to the cell-penetrating activity of the Cys2-His2 zinc-finger domain. Here, we demonstrate that genetically(More)