I am trying to design sgRNAs for dCas9 fluorescent tagging, but all of the tools I find online are for standard CRISPR knocking in/out. Anyone have a recommended tool for dCas9?
I think i used to desinged manually. just get the dna sequence complementry to the sequence where you want to make a cut. then add the restriction enzyme site to make cut for Sticky ends at both side with two different restriction enzymes, try to make the lenth under the range mentioned for guidesequence
.Then choose those enzymes which is present in your Case9 Spacer RNA or it may also be mentioned in the protocol aside. Analyzed the insertion insilico with MOLBIOTOOL restriction analyzer and then odered it. also order primers for the detection of your desired insertion so you can validate the insertion.