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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Genome editing with CRISPR–Cas nucleases, base editors, transposases and prime editors
Andrew V. Anzalone,Luke W. Koblan,Luke W. Koblan,Luke W. Koblan,David R. Liu,David R. Liu,David R. Liu +6 more
TL;DR: This work analyzes key considerations when choosing genome editing agents and identifies opportunities for future improvements and applications in basic research and therapeutics.
Therapeutic genome editing: prospects and challenges
TL;DR: Current progress toward developing programmable nuclease–based therapies as well as future prospects and challenges are discussed.
Journal ArticleDOI
Prime genome editing in rice and wheat
Qiupeng Lin,Yuan Zong,Chenxiao Xue,Shengxing Wang,Shuai Jin,Zixu Zhu,Yanpeng Wang,Andrew V. Anzalone,Andrew V. Anzalone,Andrew V. Anzalone,Aditya Raguram,Aditya Raguram,Aditya Raguram,Jordan L. Doman,Jordan L. Doman,Jordan L. Doman,David R. Liu,David R. Liu,David R. Liu,Caixia Gao +19 more
TL;DR: The resulting suite of plant prime editors enable point mutations, insertions and deletions in rice and wheat protoplasts through codon, promoter, and editing-condition optimization.
Journal ArticleDOI
The promise and challenge of therapeutic genome editing
TL;DR: The scientific, technical and ethical aspects of using CRISPR technology for therapeutic applications in humans are discussed, highlighting both opportunities and challenges of this technology to treat, cure and prevent genetic disease.
Journal ArticleDOI
Applications of CRISPR–Cas in agriculture and plant biotechnology
Haocheng Zhu,Chao Li,Caixia Gao +2 more
TL;DR: The most important applications of CRISPR-Cas in increasing plant yield, quality, disease resistance and herbicide resistance, breeding and accelerated domestication, and prospective applications of this game-changing technology are discussed.
References
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Continuous evolution of base editors with expanded target compatibility and improved activity
Benjamin W. Thuronyi,Benjamin W. Thuronyi,Benjamin W. Thuronyi,Luke W. Koblan,Luke W. Koblan,Jonathan M. Levy,Jonathan M. Levy,Wei-Hsi Yeh,Wei-Hsi Yeh,Christine R. Zheng,Gregory A. Newby,Gregory A. Newby,Christine D. Wilson,Christine D. Wilson,Mantu Bhaumik,Olga Shubina-Oleinik,Jeffrey R. Holt,David R. Liu,David R. Liu,David R. Liu +19 more
TL;DR: Phage-assisted continuous evolution of base editors (BE–PACE) is developed to improve their editing efficiency and target sequence compatibility and data from evolved CBEs are used to illuminate the relationship between deaminase activity, base editing efficiency, editing window width and byproduct formation.
Journal ArticleDOI
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Ahmed H. Badran,Ahmed H. Badran,Victor M. Guzov,Qing Huai,Melissa Kemp,Prashanth Vishwanath,Wendy Kain,Autumn M. Nance,Artem G. Evdokimov,Farhad Moshiri,Keith H. Turner,Ping Wang,Thomas M. Malvar,David R. Liu,David R. Liu +14 more
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Journal ArticleDOI
A Novel Protective Prion Protein Variant that Colocalizes with Kuru Exposure
Simon Mead,Jerome Whitfield,Mark Poulter,Paresh Shah,James Uphill,Tracy Campbell,Huda Al-Dujaily,Holger Hummerich,Jon Beck,Charles A. Mein,Claudio J. Verzilli,John C. Whittaker,Michael P. Alpers,John Collinge +13 more
TL;DR: Variants at codons 127 and 129 of PRNP demonstrate the population genetic response to an epidemic of prion disease and represent a powerful episode of recent selection in humans.
Journal ArticleDOI
A naturally occurring variant of the human prion protein completely prevents prion disease
Emmanuel A. Asante,Michelle Smidak,Andrew Grimshaw,Richard Houghton,Andrew Tomlinson,Asif Jeelani,Tatiana Jakubcova,Shyma Hamdan,Angela Richard-Londt,Jacqueline M. Linehan,Sebastian Brandner,Michael P. Alpers,Jerome Whitfield,Simon Mead,Jonathan D. F. Wadsworth,John Collinge +15 more
TL;DR: It is demonstrated that transgenic mice expressing both variant and wild-type human PrP are completely resistant to both kuru and classical Creutzfeldt–Jakob disease (CJD) prions but can be infected with variant CJD prions, a human prion strain resulting from exposure to bovine spongiform encephalopathy prions to which the Fore were not exposed.