Format

Send to

Choose Destination
Nat Protoc. 2009;4(10):1471-501. doi: 10.1038/nprot.2009.98. Epub 2009 Sep 17.

Oligomerized pool engineering (OPEN): an 'open-source' protocol for making customized zinc-finger arrays.

Author information

1
Molecular Pathology Unit, Center for Cancer Research, and Center for Computational and Integrative Biology, Massachusetts General Hospital, Charlestown, Massachusetts, USA.

Abstract

Engineered zinc-finger nucleases (ZFNs) form the basis of a broadly applicable method for targeted, efficient modification of eukaryotic genomes. In recent work, we described OPEN (oligomerized pool engineering), an 'open-source,' combinatorial selection-based method for engineering zinc-finger arrays that function well as ZFNs. We have also shown in direct comparisons that the OPEN method has a higher success rate than previously described 'modular-assembly' methods for engineering ZFNs. OPEN selections are carried out in Escherichia coli using a bacterial two-hybrid system and do not require specialized equipment. Here we provide a detailed protocol for carrying out OPEN to engineer zinc-finger arrays that have a high probability of functioning as ZFNs. Using OPEN, researchers can generate multiple, customized ZFNs in approximately 8 weeks.

PMID:
19798082
PMCID:
PMC2858690
DOI:
10.1038/nprot.2009.98
[Indexed for MEDLINE]
Free PMC Article

Supplemental Content

Full text links

Icon for Nature Publishing Group Icon for PubMed Central
Loading ...
Support Center