Broadened Cas9 cleavage
As a leading developer of CRISPR genome editing solutions, IDT continues to expand its family of Alt-R CRISPR-Cas9 nucleases with Alt-R S.p. Cas9 Nuclease V3. This CRISPR-Cas9 protein is designed to maximize the efficiency of genome editing across a broad number of sites. Alt-R S.p. Cas 9 Nuclease V3 is a recombinant variant, derived from native Streptococcus pyogenes Cas9 protein, that has been purified from an E. coli strain expressing codon optimized Cas9. Modification of the expression construct facilitates nucleus-targeted delivery, resulting in enhanced cleavage, particularly at difficult targets (Figure 1).
HiFi variant for experiments that require potent, specific editing
The IDT market-leading Alt-R HiFi Cas9 Nuclease V3 also benefits from these improvements. This high-fidelity Cas9 variant provides consistent, high editing efficiency with very low off-target activity. When delivered as part of a ribonucleoprotein (RNP) complex, other high-fidelity Cas9 enzymes have shown reduced on-target activity compared to wild-type enzymes. However, Alt-R HiFi Cas9 Nuclease V3 not only provides superior cutting specificity, but also achieves similar potency to the wide-type enzyme on the intended sites (Figure 2).
Advantages of a ribonucleoprotein delivery format
For best results, we recommend assembling Cas9 V3 nucleases with optimized Alt-R CRISPR-Cas9 crRNA and Alt-R CRISPR-Cas9 tracrRNA, and delivering them in an RNP format. Unlike conventional vector-based methods that require transcription and translation inside cells, the RNP complex can function immediately upon delivery, and is compatible with lipofection or electroporation using commercially available reagents. Moreover, the use of RNP complexes mitigates the risk of off-target cleavage by shortening the duration of Cas9 presence in cells, a major determinant of precision genome editing. Both Alt-R S.p. Cas9 Nuclease V3 and HiFi Cas9 Nuclease V3 are available in 100 and 500 µg aliquots (100 µg Cas9 nuclease = 610 pmol). Both enzymes are shipped in ready-to-use buffer at a high concentration (10 µg/µL), allowing use in a wide range of applications, including electroporation and microinjection.
Learn more about Alt-R S.p. Cas9 Nuclease V3 and Alt-R S.p. HiFi Cas9 Nuclease V3.