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CRISPRi for transcriptional repression

CRISPR interference (CRISPRi) allows researchers to down-regulate specific gene function by blocking transcription, without editing the DNA. Visit our CRISPRi applications page to learn more.

The system requires two components to operate: a gene-specific guide RNA and a catalytically deactivated Cas9 (dCas9) fused to transcriptional repressor domains (SALL1 and SDS3).

Predesigned CRISPRi guide RNA

Algorithm-designed lentiviral and synthetic single guide RNA (sgRNA) specifically target immediately downstream of the gene's transcriptional start site (TSS).

  • CRISPRi synthetic sgRNA

    The most rapid CRISPRi system, with repression within 24 hours post-transfection. Synthetic RNAs are modified for nuclease resistance and available as pooled or individual reagents.

  • CRISPRi lentiviral sgRNA

    An ideal delivery method for CRISPRi in difficult-to-transfect cells, or when longer term repression is desired.

CRISPRi dCas9-SALL1-SDS3

Nuclease-deactivated Cas9, fused to our proprietary transcriptional repressor domains SALL1 and SDS3.

CRISPRi assay considerations for reagent selection

There are many options and considerations for CRISPRi experimental conditions. Generally, users achieve the most robust knockdown when working with a stable population of dCas9-SALL1-SDS3 expressing cells. For extended timepoint assays (more than 120 hours), many choose lentiviral sgRNA. For short-term assays (less than 120 hours), synthetic sgRNA gives more robust gene repression than expressed guide RNAs. 

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

CRISPRi dCas9-SALL1-SDS3 lentiviral particles

transduction & selection for CRISPRi dCas9 stable cells

CRISPRi synthetic sgRNA Transfection or electroporation
  • Short term assays
  • Pool guide RNAs for increased repression of target gene function
  • Multiplex targets for simultaneous knockdown within gene pathways or networks
  • Arrayed screening
CRISPRi lentiviral sgRNA particles Transduction
  • Stable expression for extended timepoint assays
  • Low MOI - single integration for stable and even repression
CRISPRi dCas9-SALL1-SDS3 mRNA CRISPRi synthetic sgRNA Co-transfection or electroporation
  • Short timepoint assays - Results within hours
  • Lentiviral free workflow
  • EGFP and puromycin resistance options for enrichment of cells expressing dCas9-SALL1-SDS3 machinery
  • Pool guide RNAs for increased target gene repression
  • Multiplex targets for simultaneous knockdown within gene pathways or networks
  • Ideal for working in primary cells

CRISPRi synthetic guide RNA

CRISPRi lentiviral guide RNA

CRISPRi dCas9-SALL1-SDS3

Nuclease-deactivated Cas9, fused to our proprietary transcriptional repressor domains (SALL1 and SDS3).

CRISPRi products may be combined in several ways to support your particular experimental needs. Find workflows below for 1) co-transfection with synthetic sgRNA and dCas9-SALL1-SDS3 mRNA, 2) using CRISPRi dCas9-SALL1-SDS3 lentiviral particles to create a stable dCas9-SALL1-SDS3 expressing cell line and introducing synthetic sgRNA, or 3) creating a stable cell line and introducing either plasmid or lentiviral sgRNA

 

CRISPRi workflow using synthetic sgRNA and dCas9-SALL1-SDS3 mRNA

CRISPRi workflow with dCas9-SALL1-SDS3 mRNA and synthetic sgRNA

Co-transfect or electroporate cells with CRISPRi synthetic sgRNA and CRISPRi dCas9-SALL1-SDS3 mRNA. Then enrich cell populations using fluorescence or puromycin resistance options. This system is best suited for rapid, transient gene repression studies.


 
CRISPRi workflow using synthetic sgRNA and dCas9-SALL1-SDS3 lentiviral particles

CRISPR interference workflow with lentiviral dCas9-SALL1-SDS3 and synthetic sgRNA

Transduce cells with dCas9-SALL1-SDS3 lentiviral particles to generate CRISPRi-ready, stable dCas9-SALL1-SDS3 expressing cells. Then introduce CRISPRi synthetic sgRNA specific to your target gene(s). This system is ideal for arrayed screening in transfectable cell models.


 
CRISPRi workflow using lentiviral sgRNA and dCas9-SALL1-SDS3 lentiviral particles
CRISPRi workflow using lentiviral dCas9-SALL1-SDS3 and expressed sgRNA

Workflow using CRISPRi dCas9-SALL1-SDS3 lentiviral particles to establish a stable dCas9-SALL1-SDS3 expressing cell line, then delivering CRISPRi lentiviral sgRNA particles (left) or plasmid sgRNA (right). This system is ideal when working with difficult-to-transfect cell types.