Single Cas9 gRNA Domesticator (tS9D)

The first step for generating a CRISPR/Cas9 construct is the selection of the genomic target. The genomic target must contain a 20 nucleotides TARGET sequence followed by the PAM sequence (usually NGG). To search your genomic targets minimizing off-targeting you can use online tools such as Benchling or CRISPR-P.

Once you have selected your 20 nucleotides TARGET sequence, you can create your GB-adapted guide RNA part using the web tool below:

CRISPR Domestication
  • What's this?
  • Please, note that the target Dicot and Monocot sequences should start with 'G' and 'A' respectively. This constraint derives from the RNA polymerase III-dependent U6 and U3 promoters requirement of a 'G' or 'A' at the 5' end of the RNA to be transcribed.

  • 'The CRISPR domesticator tool will produce a domesticated sgRNA sequence flanked by standard GB overhangs (prefix and suffix). If you prefer to create your own non-standard flanking 4 nucleotide overhangs, please provide them in the boxes below (optional). Please note that if you choose the custom option the resulting oligonucleotides will not assemble with the rest of GB standard CRISPR elements.'