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Increasing the efficiency of precise genome editing with CRISPR-Cas9 by inhibition of nonhomologous end joining

Published on May 1, 2015in Nature Biotechnology 35.72
· DOI :10.1038/nbt.3190
Takeshi Maruyama13
Estimated H-index: 13
(Massachusetts Institute of Technology),
Stephanie K. Dougan25
Estimated H-index: 25
(Massachusetts Institute of Technology)
+ 3 AuthorsHidde L. Ploegh115
Estimated H-index: 115
(Massachusetts Institute of Technology)
Abstract
The efficiency of homologous recombination-based Cas9 genome editing is increased by inhibiting non-homologous end joining.
  • References (32)
  • Citations (460)
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References32
Newest
Published on Jan 1, 2015in Genetics 4.08
Priti Singh2
Estimated H-index: 2
(Cornell University),
John C. Schimenti45
Estimated H-index: 45
(Cornell University),
Ewelina Bolcun-Filas11
Estimated H-index: 11
(Cornell University)
CRISPR/Cas9 system of RNA-guided genome editing is revolutionizing genetics research in a wide spectrum of organisms. Even for the laboratory mouse, a model that has thrived under the benefits of embryonic stem (ES) cell knockout capabilities for nearly three decades, CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats)/Cas9 technology enables one to manipulate the genome with unprecedented simplicity and speed. It allows generation of null, conditional, precisely mutated, reporter...
160 Citations Source Cite
Published on Nov 1, 2014in Genome Biology 13.21
Li Song8
Estimated H-index: 8
(Johns Hopkins University),
Liliana Florea27
Estimated H-index: 27
(Johns Hopkins University),
Ben Langmead20
Estimated H-index: 20
(Johns Hopkins University)
Lighter is a fast, memory-efficient tool for correcting sequencing errors. Lighter avoids counting k-mers. Instead, it uses a pair of Bloom filters, one holding a sample of the input k-mers and the other holding k-mers likely to be correct. As long as the sampling fraction is adjusted in inverse proportion to the depth of sequencing, Bloom filter size can be held constant while maintaining near-constant accuracy. Lighter is parallelized, uses no secondary storage, and is both faster and more mem...
86 Citations Source Cite
Jiahai Shi17
Estimated H-index: 17
,
Lenka Kundrat6
Estimated H-index: 6
+ 6 AuthorsHarvey F. Lodish141
Estimated H-index: 141
(Massachusetts Institute of Technology)
We developed modified RBCs to serve as carriers for systemic delivery of a wide array of payloads. These RBCs contain modified proteins on their plasma membrane, which can be labeled in a sortase-catalyzed reaction under native conditions without inflicting damage to the target membrane or cell. Sortase accommodates a wide range of natural and synthetic payloads that allow modification of RBCs with substituents that cannot be encoded genetically. As proof of principle, we demonstrate site-specif...
72 Citations Source Cite
Published on Jun 13, 2014in Journal of Biological Chemistry 4.01
Clarissa C. Lee4
Estimated H-index: 4
(Massachusetts Institute of Technology),
Elizaveta Freinkman20
Estimated H-index: 20
(Massachusetts Institute of Technology)
+ 1 AuthorsHidde L. Ploegh115
Estimated H-index: 115
(Massachusetts Institute of Technology)
Leucine-rich repeat-containing 8 (LRRC8) proteins have been identified as putative receptors involved in lymphocyte development and adipocyte differentiation. They remain poorly characterized, and no specific function has been assigned to them. There is no consensus on how this family of proteins might function because homology searches suggest that members of the LRRC8 family act not as plasma membrane receptors, but rather as channels that mediate cell-cell signaling. Here we provide experimen...
30 Citations Source Cite
Published on Jun 1, 2014in Nature Biotechnology 35.72
Hao Yin22
Estimated H-index: 22
(Massachusetts Institute of Technology),
Wen Xue28
Estimated H-index: 28
(Massachusetts Institute of Technology)
+ 7 AuthorsDaniel G. Anderson85
Estimated H-index: 85
(Massachusetts Institute of Technology)
CRISPR-Cas9-mediated genome editing corrects a hereditary tyrosinemia disease mutation in the liver of adult mice.
512 Citations Source Cite
Published on May 1, 2014in DNA Repair 4.46
Philippe Frit6
Estimated H-index: 6
(University of Toulouse),
Nadia Barboule3
Estimated H-index: 3
(University of Toulouse)
+ 2 AuthorsPatrick Calsou10
Estimated H-index: 10
(University of Toulouse)
Abstract To cope with DNA double strand break (DSB) genotoxicity, cells have evolved two main repair pathways: homologous recombination which uses homologous DNA sequences as repair templates, and non-homologous Ku-dependent end-joining involving direct sealing of DSB ends by DNA ligase IV (Lig4). During the last two decades a third player most commonly named alternative end-joining (A-EJ) has emerged, which is defined as any Ku- or Lig4-independent end-joining process. A-EJ increasingly appears...
70 Citations Source Cite
Published on Apr 1, 2014in Nature Biotechnology 35.72
Jeffry D. Sander4
Estimated H-index: 4
(Partners HealthCare),
J. Keith Joung60
Estimated H-index: 60
The capability to introduce targeted genomic sequence changes into living cells and organisms provides a powerful tool for biological research as well as a potential avenue for therapy of genetic diseases. Frameshift knockout mutations enable reverse genetics and assignment of function, sequence insertions can be used to fuse genes to epitope tags or other functional domains, such as fluorescent proteins to endogenous gene products, and specific sequence alterations can be used to induce amino a...
1,477 Citations Source Cite
Published on Jan 3, 2014in Science 41.06
Timothy C. Wang55
Estimated H-index: 55
,
Jenny J. Wei2
Estimated H-index: 2
(Massachusetts Institute of Technology)
+ 1 AuthorsEric S. Lander245
Estimated H-index: 245
(Massachusetts Institute of Technology)
The bacterial clustered regularly interspaced short palindromic repeats (CRISPR)–Cas9 system for genome editing has greatly expanded the toolbox for mammalian genetics, enabling the rapid generation of isogenic cell lines and mice with modified alleles. Here, we describe a pooled, loss-of-function genetic screening approach suitable for both positive and negative selection that uses a genome-scale lentiviral single-guide RNA (sgRNA) library. sgRNA expression cassettes were stably integrated into...
1,311 Citations Source Cite
Published on Jan 1, 2014in Nature Reviews Molecular Cell Biology 35.61
Stephanie Panier15
Estimated H-index: 15
,
Simon J. Boulton47
Estimated H-index: 47
The function of 53BP1 in DNA double-strand break repair is multifaceted, and includes mediator and effector roles. New appreciation of how it is recruited to damaged chromatin, and how it exerts control on pathway choice, has cemented the central role of 53BP1 in genome stability maintenance.
374 Citations Source Cite
Published on Dec 1, 2013in Cell Stem Cell 23.29
Yuxuan Wu9
Estimated H-index: 9
(Chinese Academy of Sciences),
Dan Liang7
Estimated H-index: 7
(Chinese Academy of Sciences)
+ 6 AuthorsJinsong Li27
Estimated H-index: 27
(Chinese Academy of Sciences)
The CRISPR-Cas9 system has been employed to generate mutant alleles in a range of different organisms. However, so far there have not been reports of use of this system for efficient correction of a genetic disease. Here we show that mice with a dominant mutation in Crygc gene that causes cataracts could be rescued by coinjection into zygotes of Cas9 mRNA and a single-guide RNA (sgRNA) targeting the mutant allele. Correction occurred via homology-directed repair (HDR) based on an exogenously sup...
328 Citations Source Cite
Cited By460
Newest
Published on Mar 8, 2019in Nature Communications 12.35
Grégoire Cullot (French Institute of Health and Medical Research), Julian Boutin (French Institute of Health and Medical Research)+ 17 AuthorsVéronique Guyonnet-Duperat12
Estimated H-index: 12
(French Institute of Health and Medical Research)
CRISPR-Cas9 is a promising technology for genome editing. Here we use Cas9 nuclease-induced double-strand break DNA (DSB) at the UROS locus to model and correct congenital erythropoietic porphyria. We demonstrate that homology-directed repair is rare compared with NHEJ pathway leading to on-target indels and causing unwanted dysfunctional protein. Moreover, we describe unexpected chromosomal truncations resulting from only one Cas9 nuclease-induced DSB in cell lines and primary cells by a p53-de...
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Published on Mar 18, 2019
Xueli Tian (Zhengzhou University), Tingxuan Gu + 3 AuthorsZigang Dong2
Estimated H-index: 2
The development of genetic engineering in the 1970s marked a new frontier in genome-editing technology. Gene-editing technologies have provided a plethora of benefits to the life sciences. The clustered regularly interspaced short palindromic repeats/CRISPR associated protein 9 (CRISPR/ Cas9) system is a versatile technology that provides the ability to add or remove DNA in the genome in a sequence-specific manner. Serious efforts are underway to improve the efficiency of CRISPR/Cas9 targeting a...
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Published on Feb 8, 2019
Ryuichi Ono1
Estimated H-index: 1
,
Yukuto Yasuhiko9
Estimated H-index: 9
+ 3 AuthorsYoko Hirabayashi
The CRISPR-Cas9 system has been successfully applied in many organisms as a powerful genome-editing tool. Undoubtedly, it will soon be applied to human genome editing, including gene therapy. We have previously reported that unintentional DNA sequences derived from retrotransposons, genomic DNA, mRNA and vectors are captured at double-strand breaks (DSBs) sites when DSBs are introduced by the CRISPR-Cas9 system. Therefore, it is possible that unintentional insertions associated with DSB repair r...
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Published on May 23, 2019
Yagiz Aksoy1
Estimated H-index: 1
(Macquarie University),
David T. Nguyen1
Estimated H-index: 1
(Garvan Institute of Medical Research)
+ 4 AuthorsDaniel Hesselson14
Estimated H-index: 14
(University of New South Wales)
Precise genome editing is limited by the inefficiency of homology-directed repair (HDR) compared to the non-homologous end-joining (NHEJ) of double strand breaks (DSBs). The CRISPR (clustered regularly interspaced short palindromic repeat)/Cas9 system generates precise, locus-specific DSBs that can serve as substrates for HDR. We developed an in vivo visual reporter assay to quantify HDR-mediated events at single-cell resolution in zebrafish and used this system to identify small-molecule modula...
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Published on Mar 18, 2019in Scientific Reports 4.12
Sachiko Okamoto9
Estimated H-index: 9
,
Yasunori Amaishi3
Estimated H-index: 3
+ 2 AuthorsJunichi Mineno16
Estimated H-index: 16
Target-specific genome editing using engineered nucleases has become widespread in various fields. Long gene knock-in and single-base substitutions can be performed by homologous recombination (HR), but the efficiency is usually very low. To improve the efficiency of knock-in with single-stranded oligo DNA nucleotides (ssODNs), we have investigated optimal design of ssODNs in terms of the blocking mutation, orientation, size, and length of homology arms to explore the optimal parameters of ssODN...
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Published on Jul 1, 2019in Theriogenology 2.14
Ju Zhang1
Estimated H-index: 1
(Inner Mongolia University),
Jian Liu (Inner Mongolia University)+ 8 AuthorsHao Liang2
Estimated H-index: 2
(Inner Mongolia University)
Abstract The genome editors CRISPR/Cas9 (clustered regularly interspaced short palindromicrepeats/Cas9 nuclease-null) and TALENs (transcription activator-like effector nuclease) are popularly used for targeted modification of the mammalian genome. To date, few comparative studies have been carried out to investigate the differences between the use of CRISPR/Cas9 and TALENs in genome editing for goat breeding. Here, we compared CRISPR/Cas9 and TALEN technologies at multiple levels for generating ...
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