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RNA-Guided Human Genome Engineering via Cas9
Open accessAbstract
Genome Editing: Clustered regularly interspaced short palindromic repeats (CRISPR) function as part of an adaptive immune system in a range of prokaryotes: Invading phage and plasmid DNA is targeted for cleavage by complementary CRISPR RNAs (crRNAs) bound to a CRISPR-associated endonuclease (see the Perspective by van der Oost ). Cong et al. (p. 819 , published online 3 January) and Mali et al. (p. 823 , published online 3 January) adapted this defense system to function as a genome editing tool in eukaryotic cells.
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“Upon target recognition, the DNA strands are opened in an R-loop and individually cleaved, which results in a double-strand break (DSB)22,23”
From One-step generation of tumor models by base editor multiplexing in adult stem cell-derived organoids (2023) · cites this paper as [23]
Experimental — accuracy not yet measuredThe source backs part of the claim, not all of it — or backs it for a different population, condition or comparison.
In the citing paper
“The technique also allows DNA sequence changes in pluripotent embryonic stem cells (4) that can then be cultured to produce specific tissues, such as cardiomyocytes or neurons (5).”
From A prudent path forward for genomic engineering and germline gene modification (2015) · cites this paper as [4]
Experimental — accuracy not yet measuredThe source backs part of the claim, not all of it — or backs it for a different population, condition or comparison.
In the citing paper
“In January 2013, almost simultaneously, five research articles authored by different research teams appeared, all reporting that they had achieved the goal.”
From CRISPR–Cas9: A History of Its Discovery and Ethical Considerations of Its Use in Genome Editing (2022) · cites this paper as [34]
Experimental — accuracy not yet measuredThe cited source says what the claim attributes to it.
In the citing paper
“Mali et al. created a novel version of the genome-editing tool applicable to mammalian cells, termed the CRISPR/Cas9 system, which is based on modifications of the Streptococcus pyogenes type II CRISPR system in crRNA fused to trans-encoded tracrRNA 13 .”
From Harnessing the CRISPR/Cas9 system to disrupt latent HIV-1 provirus (2013) · cites this paper as [13]
The cited source says what the claim attributes to it.
In the citing paper
“The pigs were engineered to carry 69 genomic edits, using the clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated protein 9 (Cas9)-mediated nonhomologous end joining and homology-directed repair30,31, and recombinase-mediated cassette exchange32 (Fig. 2a, Extended Data Fig. 1a and Supplementary Table 2).”
From Design and testing of a humanized porcine donor for xenotransplantation (2023) · cites this paper as [31]
In the citing paper
“When a double-stranded break is introduced into genomic DNA, cells can repair the break by using a DNA molecule with high sequence similarity as a template. If an artificial template is provided while the DNA break is introduced, this pathway provides a mean to incorporate desired DNA sequences into the genome34,35”
From CRISPR-based functional genomics for neurological disease (2020) · cites this paper as [35]
In the citing paper
“The discovery, made less than 10 years ago, that a simple two-component system of a Cas protein (Streptococcus pyogenes Cas9) bound to a single guide RNA (sgRNA) can function as a programmable nuclease33 that can be deployed to cleave specific sequences in mammalian genomes34,35 has been transformative for biomedical research and the development of gene therapies.”
From CRISPR-based functional genomics for neurological disease (2020) · cites this paper as [35]
In the citing paper
“Recently, the CRISPR/Cas system has been successfully applied to human cells19,20,21,22 and several model organisms for genome editing.23,24,25,26”
From The CRISPR/Cas9 system facilitates clearance of the intrahepatic HBV templates in vivo (2014) · cites this paper as [19]
In the citing paper
“By bioinformatic screening and efficiency/off-target prediction (14, 15), we identified four gRNA targets (protospacers; LTRs A-D) that avoid conserved transcription factor binding sites, minimizing the likelihood of altering host gene expression”
From RNA-directed gene editing specifically eradicates latent and prevents new HIV-1 infection (2014) · cites this paper as [15]
In the citing paper
“Compared with well-known, site-specific genome editing technologies, such as ZFNs and TALENs, the new CRISPR-Cas approach is more convenient and efficient [10], [12].”
From High-efficiency targeted editing of large viral genomes by RNA-guided nucleases (2014) · cites this paper as [10]
In the citing paper
“Clustered regularly interspaced short palindromic repeat (CRISPR)/Cas9 is a naturally occurring bacterial immune system that uses a novel nuclease system to protect bacteria from phage infection (6-9), and it has recently been harnessed for a variety of genome-engineering applications (10)(11)(12)(13)(14)(15)(16)(17).”
From RNA-guided endonuclease provides a therapeutic strategy to cure latent herpesviridae infection (2014) · cites this paper as [13]
In the citing paper
“Moreover, no significant cytotoxicity was observed [87,88].”
From Newer Gene Editing Technologies toward HIV Gene Therapy (2013) · cites this paper as [88]
In the citing paper
“although Cas9 protein naturally induces double stranded DNA breaks by cutting each strand with two separate motifs, a mutation within one of the enzymatic motifs of Cas9 converted it into a valuable nickase with the capability to induce homologous recombination”
From Newer Gene Editing Technologies toward HIV Gene Therapy (2013) · cites this paper as [88]
In the citing paper
“and the mutant is much safer to use with less off-target effects [88].”
From Newer Gene Editing Technologies toward HIV Gene Therapy (2013) · cites this paper as [88]
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Metadata
- Authors
- Mali, P., Yang, L., Esvelt, K. M., Aach, J., Guell, M., DiCarlo, J. E., Norville, J. E., & Church, G. M.
- Affiliations
- Harvard University (US)
- Department of Genetics, Harvard Medical School, Boston, MA 02115, USA.
- Boston University (US)
- Corresponding author
- Mali, Prashant
- Yang, Luhan
- Church, George M.
- Venue
- Science
- Published
- 2013-02-15
- Volume
- 339
- Issue
- 6121
- Pages
- 823-826
- ISSN
- 0036-8075 (ISSN-L)
- 1095-9203
- PMC
- PMC3712628
- OpenAlex
- W2003171404
- Cited by
- 9,210
- References
- 46
- Citations by year
- 2026: 286 · 2025: 408 · 2024: 518 · 2023: 568 · 2022: 727 · 2021: 789 · 2020: 795 · 2019: 813 · 2018: 864 · 2017: 863 · 2016: 884 · 2015: 697 · 2014: 680 · 2013: 307
- FWCI
- 383.86
- Type
- journal article
- Language
- en
- Open access
- green
- europepmc.org
- Locations
- Science (publishedVersion)
- PubMed (publishedVersion)
- Europe PMC (PubMed Central) (acceptedVersion)
- PubMed Central (acceptedVersion)
- RECERCAT (Consorci de Serveis Universitaris de Catalunya) (acceptedVersion)
- Repositori digital de la UPF (Universitat Pompeu Fabra) (submittedVersion)
- Funders
- National Human Genome Research Institute (P50HG005550)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › CRISPR and Genetic Engineering
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA regulation and disease
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA Interference and Gene Delivery
- Keywords
- CRISPR/Cas9, guide RNA, human genome editing, AAVS1, multiplex genome editing, induced pluripotent stem cells, K562 cells, HEK293T cells, RNA-guided nucleases
- MeSH terms
- Chromosomes, Human, Pair 19, Codon, Exons, Genetic Engineering, Humans, RNA, Genome, Human, Gene Targeting, K562 Cells, DNA Cleavage, Inverted Repeat Sequences, Genetic Loci, Induced Pluripotent Stem Cells, Clustered Regularly Interspaced Short Palindromic Repeats
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