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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High-throughput functional variant screens via in vivo production of single-stranded DNA.
Max G. Schubert,Daniel B. Goodman,Timothy M. Wannier,Divjot Kaur,Fahim Farzadfard,Timothy K. Lu,Seth L. Shipman,George M. Church +7 more
TL;DR: The retron library recombineering (RLR) as discussed by the authors is a methodology for high-throughput functional screens that surpasses the scale and specificity of CRISPR-Cas methods.
Journal ArticleDOI
Applications of CRISPR/Cas to Improve Crop Disease Resistance: Beyond Inactivation of Susceptibility Factors
Dirk Schenke,Daguang Cai +1 more
TL;DR: It is concluded that genome editing by CRISPR/Cas will become increasingly indispensable to generate in relatively short time beneficial resistance traits in crops to meet upcoming challenges.
Journal ArticleDOI
Kidney Organoids and Tubuloids.
Fjodor A. Yousef Yengej,Fjodor A. Yousef Yengej,Jitske Jansen,Jitske Jansen,Maarten B. Rookmaaker,Marianne C. Verhaar,Hans Clevers +6 more
TL;DR: The nature of kidney organoids and tubuloids and their current and future applications in science and medicine are discussed and it is suggested that both organoid types hold great potential for regenerative medicine.
Journal ArticleDOI
Genomic and functional genomics analyses of gluten proteins and prospect for simultaneous improvement of end-use and health-related traits in wheat.
TL;DR: The genomic and functional genomics information obtained in the last 10 years on gluten protein chromosome loci and genes and the cis - and trans -factors regulating their expression in the grains are summarized, as well as the efforts in elucidating the involvement of gluten proteins in several wheat sensitivities affecting genetically susceptible human individuals.
Journal ArticleDOI
Efficient in vivo base editing via single adeno-associated viruses with size-optimized genomes encoding compact adenine base editors
Jessie Rose Davis,Xiao Wang,Isaac P. Witte,Tony P. Huang,Jonathan M. Levy,Aditya Raguram,Samagya Banskota,Nabil G. Seidah,Kiran Musunuru,David R. Liu +9 more
TL;DR: In this article , single-AAV-encoded ABEs retroorbitally injected in mice led to editing efficiencies in liver (66%), heart (33%), and muscle (22%) tissues that were up to 2.5-fold those of dual-AAV ABE8e, and to a 93% knockdown (on average) of human PCSK9 and of mouse Pcsk9 and Angptl3 in circulation, concomitant with substantial reductions of plasma cholesterol and triglycerides.
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
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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
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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
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