Multiplexed Cas9 targeting reveals genomic location effects and gRNA-based staggered breaks influencing mutation efficiency
نویسندگان
چکیده
منابع مشابه
Precise Cas9 targeting enables genomic mutation prevention.
Here, we present a generalized method of guide RNA "tuning" that enables Cas9 to discriminate between two target sites that differ by a single-nucleotide polymorphism. We employ our methodology to generate an in vivo mutation prevention system in which Cas9 actively restricts the occurrence of undesired gain-of-function mutations within a population of engineered organisms. We further demonstra...
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To maximise the efficiency of template-dependent gene editing, most studies describe programmable and/or RNA-guided endonucleases that make a double-stranded break at, or close to, the target sequence to be modified. The rationale for this design strategy is that most gene repair tracts will be very short. Here, we describe a CRISPR Cas9/gRNA selection-free strategy which uses deep sequencing t...
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The CRISPR/Cas or Cas9/guide RNA system is a newly developed, easily engineered and highly effective tool for gene targeting; it has considerable off-target effects in cultured human cells and in several organisms. However, the Cas9/guide RNA target site is too short for existing alignment tools to exhaustively and effectively identify potential off-target sites. CasOT is a local tool designed ...
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The relative ease of using Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)associated proteins (Cas-proteins) in genome editing has rapidly made this RNA-guided endonuclease system the method of choice in almost all current applications involving genetic manipulations, with Cas9 from S. pyogenes (SpCas9) being the most well characterized and broadly used [1]. Compared to the e...
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ژورنال
عنوان ژورنال: Nature Communications
سال: 2019
ISSN: 2041-1723
DOI: 10.1038/s41467-019-09551-w