Journal
NUCLEIC ACIDS RESEARCH
Volume 42, Issue 6, Pages -Publisher
OXFORD UNIV PRESS
DOI: 10.1093/nar/gkt1326
Keywords
-
Categories
Funding
- National Institutes of Health (NIH Nanomedicine Development Center Award) [PN2EY018244]
- National Science Foundation Graduate Research Fellowship [DGE-1148903]
- National Institutes of Health
Ask authors/readers for more resources
Although engineered nucleases can efficiently cleave intracellular DNA at desired target sites, major concerns remain on potential 'off-target' cleavage that may occur throughout the genome. We developed an online tool: predicted report of genome-wide nuclease off-target sites (PROGNOS) that effectively identifies off-target sites. The initial bioinformatics algorithms in PROGNOS were validated by predicting 44 of 65 previously confirmed off-target sites, and by uncovering a new off-target site for the extensively studied zinc finger nucleases (ZFNs) targeting C-C chemokine receptor type 5. Using PROGNOS, we rapidly interrogated 128 potential off-target sites for newly designed transcription activator-like effector nucleases containing either Asn-Asn (NN) or Asn-Lys (NK) repeat variable di-residues (RVDs) and 3- and 4-finger ZFNs, and validated 13 bona fide off-target sites for these nucleases by DNA sequencing. The PROGNOS algorithms were further refined by incorporating additional features of nuclease-DNA interactions and the newly confirmed off-target sites into the training set, which increased the percentage of bona fide off-target sites found within the top PROGNOS rankings. By identifying potential off-target sites in silico, PROGNOS allows the selection of more specific target sites and aids the identification of bona fide off-target sites, significantly facilitating the design of engineered nucleases for genome editing applications.
Authors
I am an author on this paper
Click your name to claim this paper and add it to your profile.
Reviews
Recommended
No Data Available