Patent
Cas9-dna targeting unit chimeras
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TLDR
In this article, a Cas9-DTU fusion protein was proposed to facilitate single-site nuclease gene editing precision within a human genome. But, this method is not suitable for single-genomic-site accuracy.Abstract:
The present invention provides a Cas9 platform to facilitate single-site nuclease gene editing precision within a human genome. For example, a Cas9 nuclease/DN A- targeting unit (Cas9-DTU) fusion protein precisely delivers a Cas9/sgRNA complex to a specific target site within the genome for subsequent sgRNA-dependent cleavage of an adjacent target sequence. Alternatively, attenuating Cas9 binding using mutations to the a protospacer adjacent motif (PAM) recognition domain makes Cas9 target site recognition dependent on the associated DTU, all while retaining Cas9's sgRNA-mediated DNA cleavage fidelity. Cas9-DTU fusion proteins have improved target site binding precision, greater nuclease activity, and a broader sequence targeting range than standard Cas9 systems. Existing Cas9 or sgRNA variants (e.g., truncated sgRNAs (tru-gRNAs), nickases and Fokl fusions) are compatible with these improvements to further reduce off-target cleavage. A robust, broadly applicable strategy is disclosed to impart Cas9 genome-editing systems with the single-genomic-site accuracy needed for safe, effective clinical application.read more
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References
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Journal ArticleDOI
Amino acid–base interactions: a three-dimensional analysis of protein–DNA interactions at an atomic level
TL;DR: The majority of amino acid-base interactions observed follow general principles that apply across all protein-DNA complexes, although there are individual exceptions.
Journal ArticleDOI
Genome editing with Cas9 in adult mice corrects a disease mutation and phenotype
Hao Yin,Wen Xue,Sidi Chen,Roman L. Bogorad,Eric Benedetti,Markus Grompe,Victor Koteliansky,Phillip A. Sharp,Tyler Jacks,Daniel G. Anderson +9 more
TL;DR: In this article, the authors demonstrate CRISPR-Cas9-mediated correction of a Fah mutation in hepatocytes in a mouse model of the human disease hereditary tyrosinemia.
Journal ArticleDOI
Fusion of catalytically inactive Cas9 to FokI nuclease improves the specificity of genome modification
TL;DR: In human cells, fCas9 modified target DNA sites with >140-fold higher specificity than wild-type Cas9 and with an efficiency similar to that of paired Cas9 'nickases', recently engineered variants that cleave only one DNA strand per monomer.
Journal ArticleDOI
Gene Therapy Using Adeno-Associated Virus Vectors
Shyam Daya,Kenneth I. Berns +1 more
TL;DR: Several novel approaches and recent findings that promise to expand AAV's utility are discussed, especially in the context of combining gene therapy ex vivo with new advances in stem or progenitor cell biology.
Journal ArticleDOI
Controlling signal transduction with synthetic ligands
TL;DR: Cell permeable, synthetic ligands were devised that can be used to control the intracellular oligomerization of specific proteins to demonstrate their utility and have the potential to be applied wherever precise control of a signal transduction pathway is desired.
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