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CRISPR Interference

Repress gene expression without cutting the DNA

CRISPR interference (CRISPRi) allows precise, reversible down-regulation of specific gene function by blocking transcription, without permanently editing the genome. Unlike traditional CRISPR-Cas9 knockout approaches that introduce double-strand breaks, CRISPRi uses a catalytically inactive Cas9 (dCas9) fused to transcriptional repressor domains to sterically block RNA polymerase and silence target genes at the transcriptional level.

Our CRISPRi reagents provide a straightforward, efficient toolkit for studying gene function, validating therapeutic targets, and dissecting complex biological pathways. Featuring our proprietary SALL1-SDS3 repressor system, these reagents deliver robust, tunable knockdown with minimal off-target effects, ideal for applications requiring reversible gene silencing, dose-dependent repression, or multiplexed perturbation screens.

CRISPRi is particularly valuable for essential genes, non-coding RNAs, and regulatory elements where complete knockout would be lethal or uninformative. Visit our CRISPRi applications page to explore how this cutting-edge technology can accelerate your research.

Image representing CRISPR interference

The Dharmacon™ DyadGuide CRISPR system

Reagents

CRISPRi guide RNA

CRISPRi dCas9-SALL1-SDS3

Controls, additional reagents, and libraries

CRISPR interference (CRISPRi) is a robust method for specific gene knockdown.

CRISPRi benefits

  • CRISPR specificity

    PAM-anchored targeting system

  • Gentle knockdown

    Knock genes down, not out

  • Extended time course

    Longer repression duration compared to siRNA

  • Down-regulation

    Within the gene's native context

Ideal applications

  • Loss-of-function

    Study gene function in biologically relevant knockdown models

  • Multiplexing

    Well-suited for simultaneous knockdown of multiple genes

  • Orthogonal validation

    Use in parallel with CRISPRko and/or RNAi for robust data

CRISPRi reagent selection

There are many options and considerations for CRISPRi experimental conditions. For extended timepoint assays (more than 120 hours), we recommend lentiviral sgRNA formats. For short-term assays, synthetic sgRNA typically provides more robust gene repression.

 

CRISPRi dCas9-SALL1-SDS3 source Guide RNA format Delivery method Benefits & Recommended uses

CRISPRi dCas9-SALL1-SDS3 lentiviral particle
 transduction & selection for CRISPRi dCas9 stable cells

 CRISPRi synthetic sgRNA Transfection or electroporation
  • Transient repression assays
  • Pool guide RNAs for increased repression
  • Multiplex targets for simultaneous knockdown 
  • Arrayed screening
CRISPRi lentiviral sgRNA Transduction
  • Stable expression for extended repression
  • Low MOI - single integration for stable and even repression
CRISPRi dCas9-SALL1-SDS3 mRNA CRISPRi synthetic sgRNA Co-transfection or electroporation
  • Rapid transient repression - Results within hours
  • Lentiviral free workflow
  • EGFP and puromycin resistance options for enrichment of dCas9-expressing cells
  • Pool guide RNAs for increased repression
  • Multiplex targets for simultaneous knockdown 
  • Ideal for working in primary cells
Strict-R inducible CRISPRi lentiviral system CRISPRi lentiviral sgRNA Transduction
  • Achieve reversible, small molecule-induced repression of target genes with the inducible CRISPRi Tet-Degron system, offering tight and tunable control of gene expression.
  • The system enables dCas9-SALL1-SDS3 expression upon the addition of doxycycline and Shield1, allowing dynamic, time-controlled studies of gene silencing.
CRISPRi all-in-one dCas9-SALL1-SDS3 + sgRNA Transduction
  • Maximize successful delivery of CRISPRi components into difficult-to-transduce cell types
  • Puromycin or GFP markers allow for live cell tracking of gene interference
CRISPRi lentiviral and All-in-one lentiviral dual-sgRNA Transduction
  • Deliver two sgRNAs targeting either the same gene or two different genes in a single lentiviral vector
  • Lentiviral sgRNA and All-in-one lentiviral sgRNA formats available

 

What is CRISPR interference | short video

Learn more about the Dharmacon CRISPRi system

Visit our CRISPRi applications page for more details on our novel dCas9-SALL1-SDS3 repressor construct, guide RNA design recommendations, proof-of-concept experiments and more.

Application with CRISPRi mRNA and synthetic sgRNA

See our application note on efficient multigene targeting in primary T cells with CRISPRa, CRISPRi mRNA, and synthetic guide RNAs, demonstrating the utility and feasibility of targeting multiple genes simultaneously with CRISPRi in primary T cells.


CRISPRi workflows

We offer multiple workflow options when using CRISPRi for transcriptional repression. See figure parts A, B, and C below. A and B start with transduction of dCas9-SALL1-SDS3 lentiviral particles to generate CRISPRi-ready, stable dCas9-SALL1-SDS3 expressing cells. Then sgRNAs are introduced to the stable line either as synthetic (figure part A) or lentiviral (figure part B) sgRNA to knockdown your target gene(s). For lentiviral delivery, the sgRNA and dCas9-SALL1-SDS3 can also be delivered via a single transduction in an All-in-one lentiviral format. These options are ideal for screening, extended time point assays or when working with difficult-to-transfect cell types.

Alternatively, one can (figure part C) co-transfect or electroporate dCas9-SALL1-SDS3 mRNA with synthetic sgRNA, and then enrich cell populations using fluorescence or puromycin resistance options. This system is best suited for rapid, transient gene repression studies.

CRISPRi Workflows image

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