Variant-specific knockdown by siRNA

How to use the siDESIGN Center to design isoform or gene variant-specific siRNAs

As our knowledge of the transcriptome has become more sophisticated, so has our experimental design. Dharmacon catalog siRNAs are designed to give complete knockdown by targeting all transcripts of a gene, but what options are available for a researcher that needs to target just one specific transcript variant? The siDESIGN Center offers a tool to confidently create custom, variant-specific siRNAs, by leveraging the Dharmacon proprietary siRNA design algorithm and RNA synthesis expertise with NCBI’s genomic database of information. Here are some considerations and a step-by-step guide for creating an isoform-specific siRNA for your gene of interest.

Considerations

  • Target transcript must have a stretch of sequence that is unique from the non-targeted isoforms
  • The NCBI accession number or gene ID of the targeted isoform is required

To design your isoform-specific siRNA

  1. Starting at the siDESIGN Center page, enter the gene identifier information for your target of interest (e.g.: Accession NM_005211 for human CSF1R) and press the “Advanced Options” button. Once related NCBI RefSeq accession information appears, use the radio buttons to choose the targets of interest, and exclude unwanted transcripts:
    • Click "Required" for any variant that must be targeted by the designs. Please note that at least one variant must remain "Required" for the design step to proceed.
    • Click "Allowed" for any variant that will neither be required nor excluded by resulting siRNA designs.
    • Click the "Excluded" radio button for any variant whose sequence record should NOT be targeted
    siDesign Center 
  2. To maximize the number of results, check boxes to expand the design parameters to include all regions (5’UTR, ORF, and 3’UTR).
  3. siDesign Center 
  4. Accept default G/C percentage, unless otherwise required.
  5. siDesign Center 
  6. An alignment step is required when using Advanced Options, so choose the BLAST sequence database from the pull-down that matches your species of interest and click the “Design siRNA” button. If your species is not represented, please contact Technical Support for further instructions.
  7. siDesign Center 
  8. Once results have been returned, siRNA designs are ranked with the top scoring designs on the top of the list. The Score represents a compilation of different factors created by the Dharmacon siRNA design algorithm as an indicator of the likelihood of successful silencing with a specific design. Further details about the data fields in this list can be found on the Custom siRNA Design Tool User Guide.

  9. siDesign Center 
  10. Select the ones you want and add them to your cart, and your custom designs are ready to be checked out. Once the order is placed, you can expect to receive your high-quality, transcript-specific siRNA within just 4 business days (for North America)!

Get started with the siDESIGN Center! »

Additional Resources

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Custom siDESIGN Center User Guide

  • View the online User Guide for help using siDESIGN Center tool.

Services

  • Use our online design tools and extensive synthesis options to create custom products specific to your application.

Custom RNA - Resources

  • Find product guides, FAQs and more.