Modern genomics needs updated tools: The power of updated library designs for CRISPR and RNAi screening.

Part 2 of the Realigning and Remapping Dharmacon Research Tools blog series

Why accurate gene annotation matters

The accuracy of RNA interference (RNAi) and CRISPR knockout, interference, and activation screens depends on selecting siRNAs and sgRNAs that optimize on-target activity, minimize off-target effects, and target functionally relevant gene regions based on updated genome annotations. Optimal library performance requires balancing on-target cutting/knockdown activity (functionality) with off-target liability (specificity). Both depend on accurate gene annotation through continuous realignment and remapping based on the latest RefSeq releases.

Revvity’s Dharmacon™ RNAi and CRISPR products combine expertise in synthetic oligonucleotide chemistry and flexible lentiviral vector systems with sophisticated, empirically validated, construct and sequence design algorithms. Critically, Dharmacon algorithms for both rational siRNA design and CRISPR-Cas9 gene editing balance specificity and functionality to deliver safe, effective solutions across a wide range of research models. Given the ongoing evolution of the genome and advancement of sequencing technologies and database curation, it is critical that RNAi and CRISPR libraries incorporate updated siRNA and sgRNA designs to reflect the latest transcript and genome annotations.

Newly redesigned libraries

For this reason, in 2026, we’ve used our proprietary CRISPR and siRNA design algorithms to deliver newly re-designed Edit-R sgRNA libraries for CRISPR knockout and ON-TARGETplus™ 2.0 siRNA libraries for RNA interference. Both are designed to include the most specific and relevant sequences for achieving efficient on-target editing/knockdown, while minimizing cross- or off-targeting events.

To highlight the importance of updated design tools in functional genomics screening, Drepanos et al. recently reported in Cell Genomics on the design of Jacquere, a CRISPR-Cas9 knockout genome-wide library with improved guide efficiency.¹ The sgRNA library targets a comprehensive set of genes identified by the RefSeq, GENCODE, and CHESS databases, referring to the primary transcript in all cases. This approach is consistent with that used for the Dharmacon Edit-R sgRNA libraries, which incorporate the latest RefSeq releases from late 2025 and early 2026 to update guide design.

Screening of the Jacquere library in A375 and A549 cell models showed reduced false negative rates and strong overall performance in stringent negative selection (viability) screens. These findings highlight the importance of combining updated genome annotations with powerful reagent design tools to improve the accuracy and reliability of functional genomics screens.

How we can help you

For more information on how Dharmacon screening libraries can support your genomics research, see Part 1 of the Realigning and Remapping Dharmacon Research Tools series, along with more detailed information on the importance of updating research tool designs with the ongoing evolution of the genome: for siRNA, ON-TARGETplus 2.0: Advancing siRNA design for today’s transcriptome and a recent webinar; and for CRISPR Cas9 sgRNA, Enhance Your Edits: How Our Newly Realigned Edit-R Guide RNAs Can Take Your Research Further.