Select D. melanogaster Off-Target Threshold:
Select Orthologous Gene Match Length Threshold:
You can use this tool to find Drosophila species fosmid clones suitable for cross-species RNAi rescue (Kondo et al., 2009). Fosmid clones need to be retrofitted with appropriate selection markers before use. The retrofitting vectors described in Kondo et al. (2009) are available from the DRSC.
- D. persimilis clones are available from the DRSC.
- D. pseudoobscura clones are available from BACPAC.
- D. virilis clones are available from DGRC.
For successful RNAi rescue, it is also important to use a dsRNA that does not suppress the rescue transgenes derived from a Drosophila species. The tool compares dsRNAs versus orthologs and displays the number of matching sequences longer than a user-specified threshold. This will help you choose an optimal DRSC dsRNA that has few sequence matches in the ortholog. The number of potential off-targets is also provided for each dsRNA.
If existing DRSC dsRNAs have many matching sequences to an ortholog, more suitable dsRNAs can be designed as follows: First, identify regions without long matches by comparing the cDNA sequences of D. melanogaster and a sibling species using BLAST2. Then, paste the sequence of a region without matches into SnapDragon to design optimal primers.