Related Experiment Video
Updated: Apr 16, 2026
11:37
Using Sniper-Cas9 to Minimize Off-target Effects of CRISPR-Cas9 Without the Loss of On-target Activity Via Directed Evolution
Published on: February 26, 2019
10.5K
Target specificity of the CRISPR-Cas9 system
Xuebing Wu1, Andrea J Kriz2, Phillip A Sharp3
1David H. Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, MA 02139, USA ; Computational and Systems Biology Graduate Program, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
Quantitative Biology (Beijing, China)
|February 28, 2015
Summary
The CRISPR-Cas9 system offers powerful genome editing but can cause unintended DNA alterations. This review examines the target specificity of CRISPR-Cas9, a critical factor for its therapeutic applications.
Area of Science:
- Molecular Biology
- Genetics
- Biotechnology
Background:
- The CRISPR-Cas9 system, originally a prokaryotic defense mechanism, is now a versatile RNA-guided DNA targeting tool.
- It is extensively utilized for genome editing and modulating gene expression.
- Significant potential exists for correcting mutations in human genetic diseases.
Purpose of the Study:
- To review the current understanding of CRISPR-Cas9 system's target specificity.
- To highlight the critical issue of off-target effects in CRISPR-Cas9 applications.
Main Methods:
- Literature review of studies on CRISPR-Cas9 target specificity.
- Analysis of reported off-target effects and their mechanisms.
- Evaluation of strategies to enhance CRISPR-Cas9 specificity.
Main Results:
- CRISPR-Cas9 exhibits high on-target activity but can also lead to unintended modifications at similar DNA sequences.
- Off-target effects are influenced by factors such as guide RNA design, Cas9 enzyme variant, and target DNA accessibility.
- Various methods, including modified guide RNAs and engineered Cas9 nucleases, are being developed to improve specificity.
Conclusions:
- Improving the target specificity of the CRISPR-Cas9 system is paramount for safe and effective therapeutic applications.
- Continued research into the factors influencing CRISPR-Cas9 specificity is essential for minimizing off-target effects.
- Enhanced specificity will broaden the potential of CRISPR-Cas9 in treating genetic disorders.
Related Concept Videos
01:28CRISPR/Cas9 Genome Editing
3.2K
The CRISPR-Cas system serves as a bacterial defense mechanism against invading genetic elements such as viruses and plasmids, forming the foundation for its adaptation as a powerful genome-editing tool. Originally discovered in prokaryotes, this system has been repurposed to revolutionize genetic engineering across a wide range of organisms, including plants, animals, and humans. The core component, Cas9, is an endonuclease derived from Streptococcus pyogenes, capable of introducing...
3.2K
01:59CRISPR
60.3K
Genome editing technologies allow scientists to modify an organism’s DNA via the addition, removal, or rearrangement of genetic material at specific genomic locations. These types of techniques could potentially be used to cure genetic disorders such as hemophilia and sickle cell anemia. One popular and widely used DNA-editing research tool that could lead to safe and effective cures for genetic disorders is the CRISPR-Cas9 system. CRISPR-Cas9 stands for Clustered Regularly Interspaced...
60.3K
01:59CRISPR
19.0K
19.0K
02:53CRISPR and crRNAs
20.4K
Bacteria and archaea are susceptible to viral infections just like eukaryotes; therefore, they have developed a unique adaptive immune system to protect themselves. Clustered regularly interspaced short palindromic repeats and CRISPR-associated proteins (CRISPR-Cas) are present in more than 45% of known bacteria and 90% of known archaea.
The CRISPR-Cas system stores a copy of foreign DNA in the host genome and uses it to identify the foreign DNA upon reinfection. CRISPR-Cas has three different...
The CRISPR-Cas system stores a copy of foreign DNA in the host genome and uses it to identify the foreign DNA upon reinfection. CRISPR-Cas has three different...
20.4K
02:31Homologous Recombination
65.9K
The basic reaction of homologous recombination (HR) involves two chromatids that contain DNA sequences sharing a significant stretch of identity. One of these sequences uses a strand from another as a template to synthesize DNA in an enzyme-catalyzed reaction. The final product is a novel amalgamation of the two substrates. To ensure an accurate recombination of sequences, HR is restricted to the S and G2 phases of the cell cycle. At these stages, the DNA has been replicated already and the...
65.9K
02:53Conservative Site-specific Recombination and Phase Variation
7.4K
Because the DNA segments are cut and reorganized in a direction-specific manner, site-specific recombination has emerged as an efficient genetic engineering technique. Flippase and Cyclization recombinases or Flp and Cre, respectively, are two members of the tyrosine recombinase family derived from bacteriophages, that are used to mediate site-specific DNA insertions, deletions, and targeted expression of proteins in mammalian cell lines.
The recognition sites for Cre recombinase called LoxP...
The recognition sites for Cre recombinase called LoxP...
7.4K