Basic questions
What is this database all about?
See the About page for details on CRISPR/Cas9 technology and how guide RNAs were identified and selected for this database.
What is an"NGG PAM sequence" ?
This is the protospacer adjacent motif (PAM) immediately following the crRNA target sequence in the genomic DNA. Cas9 protein variants all require a unique PAM site for binding and nuclease activity. NGG is the PAM site for Cas9 from Streptococcus pyrogenes.
What is an"NGA PAM sequence" ?
This PAM site is recognized by the VQR variant of Cas9 from Streptococcus pyrogenes (Kleistiver et al, Nature. 2015).
What is "folding energy" ?
The folding energy is the result of a minimum free energy calculation in kcal/mol for each guide in the database. Users should avoid selecting guides with large negative values as they are more likely to form stable secondary structures which could reduce efficiency.
What is "GG before the PAM sequence" or "3' GG status" ?
Farboud and Meyer (Genetics, 2015) found there was increased efficiency of Cas9 cutting if the 20 base pair protospacer (crRNA) ended in GG immediately before the PAM site.
What does "web page" versus "text only" mean?
Use "web page", if you want to view the search results in your web browser. Use "text only" for a tab-delimited text-based output in a browser window that can be copied and further processed locally. The sequence names in the html output table are hyperlinks to the corresponding gene page in Wormbase. The "cut site position" is a hyperlink to the genome browser view of the location in Wormbase.
Frequently asked questions
Why are there no guide RNAs in my gene of interest?
- make sure you use the correct type of identifier (gene name or sequence name, see Wormbase for details)
- make sure you don't use CGC gene names that are not in Wormbase release WB250
- make sure your search criteria are consistent (e.g. check 'RNA' under features, if your gene encodes a non-coding RNA
Why are there no guide RNAs in my chromosomal interval?
- make sure you haven't entered anything in the gene search field