Hello genome editors,
When using the CRISPR-Cas9 system, does anyone know how long the gRNAs are usually expressed for? I know that they are generally expressed from a constitutive pol III promoter, but is there any evidence that the gRNA expression vector is itself targeted for cleavage? Does it require a PAM?
Thanks,
John
The gRNA vector itself is not targeted for cleavage since it doesn't have the PAM sequence which is also required for the Cas9 to cleave the target.
In a transient transfection system, the transgene is expression peaks by 24 hour and start to fall by 96 hours. By 7 days there is barely any expression due to loss of plasmid due to cell division and other factors.
If your gRNA expression construct contains exactly same sequence, including PAM, as your target sequence then it might get cut.
- Badges
- Science topic
- Similar topics
- Biological Science
- Immunology
- Antibodies