Search-and-replace genome editing without double-strand breaks or donor DNA
Andrew V. Anzalone,Andrew V. Anzalone,Andrew V. Anzalone,Peyton B. Randolph,Peyton B. Randolph,Peyton B. Randolph,Jessie Rose Davis,Jessie Rose Davis,Jessie Rose Davis,Alexander A. Sousa,Alexander A. Sousa,Alexander A. Sousa,Luke W. Koblan,Luke W. Koblan,Luke W. Koblan,Jonathan M. Levy,Jonathan M. Levy,Jonathan M. Levy,Peter J. Chen,Peter J. Chen,Peter J. Chen,Christine D. Wilson,Christine D. Wilson,Christine D. Wilson,Gregory A. Newby,Gregory A. Newby,Gregory A. Newby,Aditya Raguram,Aditya Raguram,Aditya Raguram,David R. Liu,David R. Liu,David R. Liu +32 more
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TLDR
A new DNA-editing technique called prime editing offers improved versatility and efficiency with reduced byproducts compared with existing techniques, and shows potential for correcting disease-associated mutations.Abstract:
Most genetic variants that contribute to disease1 are challenging to correct efficiently and without excess byproducts2-5. Here we describe prime editing, a versatile and precise genome editing method that directly writes new genetic information into a specified DNA site using a catalytically impaired Cas9 endonuclease fused to an engineered reverse transcriptase, programmed with a prime editing guide RNA (pegRNA) that both specifies the target site and encodes the desired edit. We performed more than 175 edits in human cells, including targeted insertions, deletions, and all 12 types of point mutation, without requiring double-strand breaks or donor DNA templates. We used prime editing in human cells to correct, efficiently and with few byproducts, the primary genetic causes of sickle cell disease (requiring a transversion in HBB) and Tay-Sachs disease (requiring a deletion in HEXA); to install a protective transversion in PRNP; and to insert various tags and epitopes precisely into target loci. Four human cell lines and primary post-mitotic mouse cortical neurons support prime editing with varying efficiencies. Prime editing shows higher or similar efficiency and fewer byproducts than homology-directed repair, has complementary strengths and weaknesses compared to base editing, and induces much lower off-target editing than Cas9 nuclease at known Cas9 off-target sites. Prime editing substantially expands the scope and capabilities of genome editing, and in principle could correct up to 89% of known genetic variants associated with human diseases.read more
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Fine-tuning shoot meristem size to feed the world
TL;DR: In this article , the authors highlight recent progress in understanding how shoot meristem size affects yield-related traits and outline the strategies to fine-tune shoot MERISTEM regulatory genes to meet the demands of a growing population and promote sustainable agriculture.
Journal ArticleDOI
Fine-tuning shoot meristem size to feed the world
TL;DR: In this article, the authors highlight recent progress in understanding how shoot meristem size affects yield-related traits and outline the strategies to fine-tune shoot regulatory genes to meet the demands of a growing population and promote sustainable agriculture.
Journal ArticleDOI
A primer to gene therapy: Progress, prospects, and problems.
TL;DR: The features of state‐of‐the‐art viral vector systems and the corresponding gene‐centered therapies they seek to deliver and a brief summary is given on emerging gene editing approaches built on CRISPR‐Cas9 nucleases and, more recently, nickases, including base editors and prime editors.
Journal ArticleDOI
CRISPR-Cas9-based genetic engineering for crop improvement under drought stress.
Abdul Sami,Zhao Xue,Saheera Tazein,Ayesha Arshad,Zong He Zhu,Ya Ping Chen,Yue Hong,Xiao Tian Zhu,Ke Jin Zhou +8 more
TL;DR: In this paper, the authors highlight the principle and optimization of CRISPR systems and their implementation for crop improvement, particularly in terms of drought tolerance, yield, and domestication, and address the ways in which innovative genome editing tools can help recognize and modify novel genes coffering drought tolerance.
Journal ArticleDOI
Chemical genetics strategy to profile kinase target engagement reveals role of FES in neutrophil phagocytosis
Tom van der Wel,Riet Hilhorst,Hans den Dulk,Tim van den Hooven,Nienke M. Prins,Joost A. P. M. Wijnakker,Bogdan I. Florea,Eelke B. Lenselink,Gerard J. P. van Westen,Rob Ruijtenbeek,Herman S. Overkleeft,Allard Kaptein,Tjeerd Barf,Mario van der Stelt +13 more
TL;DR: A chemical genetics strategy to study target engagement of endogenous kinases, achieving specific labeling and inactivation of FES kinase to provide insights into FES’ role in neutrophil phagocytosis holds promise as a target validation method for kinases.
References
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limma powers differential expression analyses for RNA-sequencing and microarray studies
Matthew E. Ritchie,Belinda Phipson,Di Wu,Yifang Hu,Charity W. Law,Wei Shi,Gordon K. Smyth,Gordon K. Smyth +7 more
TL;DR: The philosophy and design of the limma package is reviewed, summarizing both new and historical features, with an emphasis on recent enhancements and features that have not been previously described.
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RSEM: accurate transcript quantification from RNA-Seq data with or without a reference genome
Bo Li,Colin N. Dewey +1 more
TL;DR: It is shown that accurate gene-level abundance estimates are best obtained with large numbers of short single-end reads, and estimates of the relative frequencies of isoforms within single genes may be improved through the use of paired- end reads, depending on the number of possible splice forms for each gene.
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Martin Jinek,Krzysztof Chylinski,Krzysztof Chylinski,Ines Fonfara,Michael H. Hauer,Jennifer A. Doudna,Emmanuelle Charpentier +6 more
TL;DR: This study reveals a family of endonucleases that use dual-RNAs for site-specific DNA cleavage and highlights the potential to exploit the system for RNA-programmable genome editing.
Journal ArticleDOI
Multiplex Genome Engineering Using CRISPR/Cas Systems
Le Cong,Le Cong,F. Ann Ran,F. Ann Ran,David M. Cox,David M. Cox,Shuailiang Lin,Shuailiang Lin,Robert P. J. Barretto,Naomi Habib,Patrick D. Hsu,Patrick D. Hsu,Xuebing Wu,Wenyan Jiang,Luciano A. Marraffini,Feng Zhang +15 more
TL;DR: The type II prokaryotic CRISPR (clustered regularly interspaced short palindromic repeats)/Cas adaptive immune system has been shown to facilitate RNA-guided site-specific DNA cleavage as discussed by the authors.
Multiplex Genome Engineering Using CRISPR/Cas Systems
Le Cong,F. A. Ran,David Benjamin Turitz Cox,Shuailiang Lin,Robert P. J. Barretto,Naomi Habib,Patrick D. Hsu,Xuebing Wu,Wenyan Jiang,Luciano A. Marraffini,Feng Zhang +10 more
TL;DR: Two different type II CRISPR/Cas systems are engineered and it is demonstrated that Cas9 nucleases can be directed by short RNAs to induce precise cleavage at endogenous genomic loci in human and mouse cells, demonstrating easy programmability and wide applicability of the RNA-guided nuclease technology.