In vivo genome editing using Staphylococcus aureus Cas9
By F. Ran, Le Cong, Winston X. Yan, David A. Scott, Jonathan Gootenberg, Andrea J. Kriz, Bernd Zetsche, Ophir Shalem, Xuebing Wu, Kira S. Makarova and 3 more. 2015
2,538 citations recorded by the source corpus.
The RNA-guided endonuclease Cas9 has emerged as a versatile genome-editing platform. However, the size of the commonly used Cas9 from Streptococcus pyogenes (SpCas9) limits its utility for basic research and therapeutic applications that use the highly versatile adeno-associated virus (AAV) delivery vehicle. Here, we characterize six smaller Cas9 orthologues and show that Cas9 from Staphylococcus aureus (SaCas9) can edit the genome with efficiencies similar to those of SpCas9, while being more than 1 kilobase shorter. We packaged SaCas9 and its single guide RNA expression cassette into a single AAV vector and targeted the cholesterol regulatory gene Pcsk9 in the mouse liver. Within one week of injection, we observed >40% gene modification, accompanied by significant reductions in serum Pcsk9 and total cholesterol levels. We further assess the genome-wide targeting specificity of SaCas9 and SpCas9 using BLESS, and demonstrate that SaCas9-mediated in vivo genome editing has the potential to be efficient and specific.
Subfields: Genetics, Molecular Biology
How researchers see this paper
The researchers whose trusted networks rank it highest, strongest advocate first. Every researcher gets a different order.
In Genetics
- Eugene V. Koonin
- Feng Zhang
- Lei S. Qi
- Kira S. Makarova
- Haifeng Wang
- Marie La Russa
- Rodolphe Barrangou
- David R. Liu
In Molecular Biology
- Feng Zhang
- Jennifer A. Doudna
- Lei S. Qi
- Eugene V. Koonin
- Haifeng Wang
- Marie La Russa
- Kira S. Makarova
- Rodolphe Barrangou
Citations in the corpus
- Evolved Cas9 variants with broad PAM compatibility and high DNA specificity (2018, positive): “We first restored the catalytic residues Asp 10 and His 840 to test if xCas9 nucleases can cleave DNA even though they were evolved only for DNA binding.”
- Evaluation of off-target and on-target scoring algorithms and integration into the guide RNA selection tool CRISPOR (2016, methodology): “2 % [2, 3, 33] (Additional file 1: Table S1; [26, 28] did not report sensitivity).”
- Biology and Applications of CRISPR Systems: Harnessing Nature's Toolbox for Genome Engineering. (2016, neutral): “Recent work has focused on developing various chemical- and light-inducible Cas9 constructs to allow for greater spatiotemporal control and on employing Cas9 orthologs with different PAM sequences and smaller sizes, allowing for easier packaging in adeno-associated virus vectors (Davis et al., 2015;”
- CRISPR/Cas9 in Genome Editing and Beyond. (2016, background): “, Sa Cas9 (69), St1 Cas9, and Nm Cas9 (71)].”
- Delivering CRISPR: a review of the challenges and approaches (2018, methodology): “Groups have used of SaCas9 in AAV vectors to target the cholesterol regulatory gene Pcsk9 (Ran et al., 2015) and disrupt mutations in the DMD gene in adult mice (Nelson et al., 2016; Tabebordbar et al., 2016).”
- Engineering the Delivery System for CRISPR-based Genome Editing (2018, background): “There are also smaller Cas9 orthologs than the commonly used Streptococcus pyogenes Cas9 (SpCas9), such as Staphylococcus aureas Cas9 (SaCas9) [13].”