Activation guides are located -1000bp/+200bp from the TSS and suppression guides are located -200bp/+1000bp from the TSS.
Frequently asked questions
- What is the difference between activation and suppression guides?
- How are off-targets computed?
20bp spacer sequences were sensitively mapped genome-wide using bowtie2 and then restricted to off-target sites that contained a dCasMINI PAM directly upstream of the spacer sequence. Final on-target spacer lists were sorted based on number of putative off-targets, then distance to TSS.
- What PAMs does this tool support?
We have designed this tool to support the following six PAMs: TTTG, TTTA, TCTA, CTTG, TTCA, and TTCG. In addition to the canonical TTTR PAM (Xu et al., 2021), internal studies characterizing the PAM preference of dCasMINI have motivated us to include these additional options.
- How were genome-wide TSSs determined?
For each gene in the human genome (hg38), we selected the primary TSS as reported by the FANTOM5 collection (Noguchi et al., 2017), with supplemental TSS coverage sourced from GENCODE (Frankish A et al., 2018).
- What can I do if the a target gene I'm interested in doesn't appear?
We are always looking to expand the utility of this tool. We welcome any feedback regarding gene targets the community would find useful.
- Is this tool capable of designing guides against genomes other than human?
While we currently only support guide design against the human genome (hg38), stay tuned as we plan to add additional genomes in the future to facilitate studies in model systems.