CRISPR/Cas9 genome editing has become widely used due to its simplicity and versatility, and the CRISPR technology has been adapted for diverse applications aside from genome editing. As a leader in gene synthesis and genome editing, and through our partnership with Feng Zhang's laboratory at the Broad Institute of MIT and Harvard*, GenScript offers validated CRISPR products, services and resources to help you harness the power of CRISPR genome editing for your research.
CRISPR stands for Clustered Regularly Interspaced Short Palindromic Repeats. CRISPR sequences were originally identified in the Escherichia coli (E. coli) genome, and were found to function as part of an RNA-based adaptive immune system to target and destroy genetic parasites at the DNA level. Read More »
CRISPR-associated protein (Cas) is an endonuclease that cuts foreign DNA, allowing integration into the host genome. Cleavage only occurs when there is a protospacer adjacent motif (PAM) around the targeted sequence of the invading DNA, ensuring highly accurate targeting.
Researchers studying CRISPR have adapted it for use as a tool for genetic modification of the target host genome. CRISPR/Cas9 has recently become a popular genome editing tool, due to its simplicity and versatility.
|gRNA sequence databases||gRNA design tool|
|CRISPR gRNA constructs||CRISPR gRNA libraries|
|CRISPR-edited mammalian cell lines||Microbial gene editing|
|Purified Cas9 proteins||Genomic DNA sequencing|
Have more questions about CRISPR/Cas9? Learn more about the advantages of CRISPR and how to integrate it into your research.
Of the other gene editing technologies available, CRISPR/Cas9 has stood out for its simplicity and efficacy. The CRISPR system requires only a few simple DNA constructs to encode the gRNA and Cas9, and if knock-in is being performed, the donor template for HR. As a result, CRISPR gene editing is an approachable technique for use in any lab regardless of molecular biology expertise. The table below outlines a few of the key differences between CRISPR gene editing and other popular techniques.
Once Cas9 nucleases are guided to the target DNA and create a double strand break 3-4 bases upstream from the PAM sequences, there are two ways the double strand break (DSB) can be repaired. If there is no donor DNA present, resolution will occur by error-prone non-homologous end joining (NHEJ), resulting in an indel that effectively knocks out protein function. Alternatively, if donor DNA sequences are available, the DSB is repaired by homology directed repair (HDR) for precise knock-in of the target gene.
The most efficient method to deliver Cas9 and gRNA plasmids depends largely on the cell type. For easy-to-transfect cell lines, non-viral constructs are often suitable and can be delivered with high efficiency by lipofection. The plasmids usually contain selection markers to confirm effective delivery, such as antibiotic resistance gene s or fluorescent proteins. For hard-to-transfect cell lines, such as stem cells, viral-based transfection methods may be more effective. Lentiviral vectors may be more suitable for these cell types. All of GenScript's lentiviral vectors are compatible with 3rd and 4th generation lenti-packaging systems.
For detailed information on experimental design, we recommend consulting Ran et al's publication:
Ran et al. Genome engineering using the CRISPR-Cas9 system. Nature Protocols. 2013; 8:2281.
For more information on lentiviral transfection protocols, click here.
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