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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Evaluating CRISPR-based Prime Editing for cancer modeling and CFTR repair in intestinal organoids
Maarten H. Geurts,Eyleen de Poel,Cayetano Pleguezuelos-Manzano,Léo Carrillo,Amanda Andersson-Rolf,Matteo G Boretto,Jeffrey M. Beekman,Hans Clevers +7 more
TL;DR: Underline the broad applicability of prime editing for modeling oncogenic mutations and showcase the potential clinical application of this technique, pending further optimization, in human intestinal organoids.
Journal ArticleDOI
One-Step Homology Mediated CRISPR-Cas Editing in Zygotes for Generating Genome Edited Cattle.
Ki-Eun Park,Juli Foster Frey,Jerel J. Waters,Sean G Simpson,Chris Coutu,Sarah Plummer,Matthew Campbell,David M. Donovan,Bhanu Prakash V.L. Telugu +8 more
TL;DR: This study reports for the first time the use of CRISPR-Cas genome editors to introduce novel PRNP allelic variants that have been shown to provide resilience towards human prion pandemics.
Journal ArticleDOI
Precise genomic editing of pathogenic mutations in RBM20 rescues dilated cardiomyopathy
Takahiko Nishiyama,Yu Zhang,Miao Cui,Hui Li,Efrain Sanchez-Ortiz,John McAnally,Wei Theng Tan,Jiwoong Kim,Kenian Chen,Lin Xu,Rhonda Bassel-Duby,Eric N. Olson +11 more
TL;DR: Nishiyama et al. as mentioned in this paper used ABE and PE to correct pathogenic RBM20R636S mutations in human isogenic induced pluripotent stem cell (iPSC)-derived cardiomyocytes.
Journal ArticleDOI
De novo design of future rapeseed crops: Challenges and opportunities
TL;DR: In this article , advances in research on the physiological and genetic bases of both stress factors-affected yield stability and seed yield potential, focusing on source-sink relationships and allocation of photosynthetic assimilates to vegetative growth and seed development.
Journal ArticleDOI
Correction of the aprt Gene Using Repair-Polypurine Reverse Hoogsteen Hairpins in Mammalian Cells.
TL;DR: It is demonstrated that repair-PPRHs achieve the permanent correction of point mutations in the dsDNA at the endogenous level in mammalian cells without off-target activity.
References
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Journal ArticleDOI
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.
Journal ArticleDOI
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.
Journal ArticleDOI
A programmable dual-RNA-guided DNA endonuclease in adaptive bacterial immunity.
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.