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Can I read Quantification of Cas9 binding and cleavage across diverse guide sequences maps landscapes of target engagement on EtoBox?
Quantification of Cas9 binding and cleavage across diverse guide sequences maps landscapes of target engagement by Boyle, Evan A. (author);Becker, Winston R. (author);Bai, Hua B. (author);Chen, Janice S. (author);Doudna, Jennifer A. (author);Greenleaf, William J. (author) is a Biochemistry, Genetics and Molecular Biology article available to read on EtoBox.
What is Quantification of Cas9 binding and cleavage across diverse guide sequences maps landscapes of target engagement about?
The RNA-guided nuclease Cas9 has unlocked powerful methods for perturbing both the genome through targeted DNA cleavage and the regulome through targeted DNA binding, but limited biochemical data have hampered efforts to quantitatively model sequence perturbation of target binding and cleavage across diverse guide sequences. We present scalable, sequencing-based platforms for high-throughput filter binding and cleavage and then perform 62,444 quantitative binding and cleavage assays on 35,047 on- and off-target DNA sequences across 90 Cas9 ribonucleoproteins (RNPs) loaded with distinct guide RNAs. We observe that binding and cleavage efficacy, as well as specificity, vary substantially across RNPs; canonically studied guides often have atypically high specificity; sequence context surrounding the target modulates Cas9 on-rate; and Cas9 RNPs may sequester targets in nonproductive states that contribute to "proofreading" capability. Lastly, we distill our findings into an interpretable biophysical model that predicts changes in binding and cleavage for diverse target sequence perturbations.
Who reads Quantification of Cas9 binding and cleavage across diverse guide sequences maps landscapes of target engagement?
It is typically read by researchers, students, and practitioners in Biochemistry, Genetics and Molecular Biology.
- Author
- Boyle, Evan A. (author);Becker, Winston R. (author);Bai, Hua B. (author);Chen, Janice S. (author);Doudna, Jennifer A. (author);Greenleaf, William J. (author)
- Publisher
- American Association for the Advancement of Science (AAAS)
- Published
- 2021
- Language
- EN
- Field
- Biochemistry, Genetics and Molecular Biology (Physical Sciences)