期刊
NATURE PROTOCOLS
卷 1, 期 3, 页码 1637-1652出版社
NATURE PUBLISHING GROUP
DOI: 10.1038/nprot.2006.259
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资金
- NATIONAL CANCER INSTITUTE [T32CA009216, R21CA120681] Funding Source: NIH RePORTER
- NATIONAL INSTITUTE OF GENERAL MEDICAL SCIENCES [R01GM072621, R01GM069906] Funding Source: NIH RePORTER
- OFFICE OF THE DIRECTOR, NATIONAL INSTITUTES OF HEALTH [DP1OD006862] Funding Source: NIH RePORTER
- NCI NIH HHS [T32 CA09216, R21 CA120681] Funding Source: Medline
- NHLBI NIH HHS [R01 HL0792595] Funding Source: Medline
- NIGMS NIH HHS [R01 GM069906, R01 GM072621] Funding Source: Medline
- NIH HHS [DP1 OD006862] Funding Source: Medline
Engineered zinc finger nucleases can stimulate gene targeting at specific genomic loci in insect, plant and human cells. Although several platforms for constructing artificial zinc finger arrays using modular assembly have been described, standardized reagents and protocols that permit rapid, cross-platform mixing-and-matching of the various zinc finger modules are not available. Here we describe a comprehensive, publicly available archive of plasmids encoding more than 140 well-characterized zinc finger modules together with complementary web-based software (termed ZiFiT) for identifying potential zinc finger target sites in a gene of interest. Our reagents have been standardized on a single platform, enabling facile mixing-and-matching of modules and transfer of assembled arrays to expression vectors without the need for specialized knowledge of zinc finger sequences or complicated oligonucleotide design. We also describe a bacterial cell-based reporter assay for rapidly screening the DNA-binding activities of assembled multi-finger arrays. This protocol can be completed in approximately 24-26 d.
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