What reagents will I need in addition to the Alt-R^® RNAs for a CRISPR-Cas9 genome editing experiment?

Ribonucleoprotein (RNP) components

The Alt-R^® CRISPR-Cas9 System includes a target-specific CRISPR-Cas9 crRNA and a universal CRISPR-Cas9 tracrRNA, both of which are required for genome editing using the Alt-R CRISPR-Cas9 System protocol. Nuclease-Free Duplex Buffer, which is used to complex the crRNA and the tracrRNA, is included with the Alt-R tracrRNA. You will also need an S. pyogenes Cas9 endonuclease:

• Alt-R S.p. Cas9 Nuclease 3NLS, recommended for most experiments
• Alt-R S.p. HiFi Cas9 Nuclease 3NLS, for reduced off-target effects
• Alt-R S.p. Cas9 D10A or H840A Nickase 3NLS, for increased specificity and reduced off-target effects
• S.p. Cas9 Expression Plasmid.

Delivery

Supply your own transfection reagents to effectively deliver CRISPR reagents into your cells. We recommend delivery of an RNP for best results. The CRISPR-Cas9 System is compatible with transfection (lipid-mediated or electroporation) and microinjection.

Additional reagents

We also highly recommend the use of control crRNAs, which can be ordered as part of the Alt-R CRISPR-Cas9 Control Kits or individually.

The Alt-R Cas9 Electroporation Enhancer, which is a non-targeting carrier DNA, can increase the efficiency of electroporation. The fluorescently labeled Alt-R CRISPR-Cas9 tracrRNA – ATTO™ 550, which functions like the unlabeled Alt-R CRISPR-Cas9 tracrRNA, can be used to monitor transfection efficiency, study cellular localization, or enrich for CRISPR-edited cells.

If you are interested in additional options for genome editing, we also offer reagents to perform CRISPR-Cpf1 genome editing.