The Edit-R™ CRISPR Design Tool allows you to quickly and easily generate guide RNA sequences for MAD7 and other CRISPR-Cas systems such as Cas12a, Cas12c, Cas13, or novel CRISPR enzymes.
Our CRISPR Design Tool provides an intuitive one-stop location for guide RNA design and ordering
Genome engineering has advanced tremendously with the characterization of bacterial and archaeal CRISPR (clustered regularly interspaced short palindromic repeats) systems and their adapted usage in mammalian cells. The ability to precisely and permanently alter endogenous genomic sequence through targeted genome editing is a highly effective reverse genetics tool.
In addition to the canonical Cas9 nuclease, other editing nucleases have been discovered including Cas12a (includes MAD7), Cas12b, Cas12c, Cas13, and many more.
MAD7 is a novel nuclease that was released by Inscripta in 2017. This nuclease only requires a crRNA for gene editing and allows for specific targeting of AT rich regions of the genome. MAD7 cleaves DNA with a staggered cut as compared to S. pyogenes
which has blunt cutting.
The PAM sequence is YTTV. Y indicates a C or T base, and V indicates A, C or G. The PAM sequence is located upstream of the target sequence, and the repeat sequence appended to the 5’ of the target sequence is TTAATTTCTACTCTTGTAGAT.
The DNA cleavage sites for MAD7 relative to the target site are 19 bases after the YTTV PAM site on the sense strand and 23 bases after the ecomplementary PAM site of the anti-sense strand.
When designing experiments it is always important to include controls, the recommended crRNA control sequences for use in human cell lines are below:
These can be easily ordered through the CRISPR design tool by selecting ‘Input my own guide RNA sequence’ from the Start tab.
The following synthetic RNA products may be ordered through the CRISPR Design Tool. Contact technical support if larger quantities are needed.
Synthetic crRNA is also available at a discounted price in 96-well plates, when ordering a minimum of 20 wells.
MAD7, Cas9 or Cpf1-expressing plasmids (200 ng/well) were co-transfected with their respective synthetic guide RNAs (25 nM) into 293T cells using 0.7 µL/well of DharmaFECT Duo transfection reagent. PPIB (A) and DNMT3B (B) gene editing activities (estimated percentage of indels) were determined by a T7EI DNA mismatch detection assay.
MAD7, Cas9 proteins (25 nM) and their respective guide RNAs (50 nM) were co-transfected into HCT116 cells using 0.4 µL/well of DharmaFECT Duo transfection reagent. After 72 hours, gene editing activities (estimated percentage of indels) of PPIB (A) and STAG2 (B) were determined using T7EI DNA mismatch assay.
Design your guide RNA for CRISPR-Cas systems such as MAD7, Cas12a, Cas13a or other systems
Data that demonstrate synthetic guide RNA show efficient editing in mammalian cells
In vitro and in vivo CRISPR gene editing with MAD7