CRISPR interference reagents

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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 Dharmacon™ CRISPRmod™ CRISPRi system requires two components to operate: a gene-specific CRISPRi guide RNA and a catalytically deactivated Cas9 (dCas9) fused to transcriptional repressor domains (SALL1 and SDS3). Our CRISPRi reagents provide a straightforward, efficient set of tools to study a gene’s function via transcriptional repression.

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. 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
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 dCas9 solutions

dCas9-SALL1-SDS3 mRNA

Co-transfect with synthetic sgRNA for transient CRISPRi in lentiviral-free workflows.

dCas9-SALL1-SDS3 lentiviral particles

Create a stable dCas9 expressing cell line, ready for CRISPRi studies.

CRISPRi gene repression is observed 24 hours post-transfection and is maximal 48-72 hours post transfection — CRISPRmod CRISPRi synthetic sgRNAs achieve maximal repression at 48-72 hours post-transfection in both dCas9-KRAB and Cas9-SALL1-SDS3-expressing cells. U2OS cells stably expressing integrated dCas9-KRAB or dCas9-SALL1-SDS3 were plated at 10,000 cells/well and transfected using DharmaFECT 4 Transfection Reagent with pools of pre-designed synthetic sgRNA (25nM) targeting CBX1, HBP1, or SEL1L. Cells were harvested at 24, 48, 72, 96, 120, and 144 hours post-transfection, total RNA was isolated, and relative gene expression was measured using RT-qPCR. The relative expression of each target gene was calculated with the ∆∆Cq method using GAPDH as the housekeeping gene and normalized to a non-targeting control (NTC).

CRISPRi lentiviral sgRNA is well suited for long term repression — U2OS and A549 cells stably expressing integrated dCas9-KRAB or dCas9-SALL1-SDS3 were plated at 50,000 cells/well in 24 well plates and transduced with sgRNA lentiviral particles targeting PPIB or SEL1L at a MOI of 0.3 to obtain cells with a single integrant. Cells were selected with 2 µg/mL puromycin for 5 days, split into two populations, and allowed to recover for an additional day. At 7 days post-transduction one population was harvested for RT-qPCR analysis. The replicate populations of cells were cultured for 7 additional days before harvest. Total RNA was isolated from harvested plates and relative gene expression was calculated with the ∆∆Cq method using GAPDH as the housekeeping gene and normalized to a non-targeting control.

CRISPR interference reagents

Repress gene expression, without cutting the DNA

CRISPRi reagent selection

Order CRISPRi guide RNA

CRISPRi synthetic sgRNA

CRISPRi lentiviral sgRNA

CRISPRi All-in-one Lentiviral sgRNA

CRISPRi dCas9 solutions

dCas9-SALL1-SDS3 mRNA

dCas9-SALL1-SDS3 lentiviral particles

CRISPRi gene repression is observed 24 hours post-transfection and is maximal 48-72 hours post transfection

CRISPRi gene repression is observed 24 hours post-transfection and is maximal 48-72 hours post transfection

CRISPRi lentiviral sgRNA is well suited for long term repression

CRISPRi synthetic guide RNA

CRISPRi synthetic sgRNA

CRISPRi synthetic sgRNA positive controls

CRISPRi synthetic sgRNA non-targeting controls

CRISPRi lentiviral guide RNA

CRISPRi lentiviral sgRNA

CRISPRi lentiviral sgRNA positive controls

CRISPRi lentiviral sgRNA non-targeting controls

CRISPRi dCas9-SALL1-SDS3

CRISPRi dCas9-SALL1-SDS3 mRNA

CRISPRi dCas9-SALL1-SDS3 lentiviral particles

CRISPRi screening libraries

Custom cherry-pick libraries

Horizon's CRISPRi workflows

CRISPRi workflows

CRISPR interference reagents

Repress gene expression, without cutting the DNA

CRISPRi reagent selection

Order CRISPRi guide RNA

CRISPRi dCas9 solutions

Supporting data

CRISPRi gene repression is observed 24 hours post-transfection and is maximal 48-72 hours post transfection

CRISPRi gene repression is observed 24 hours post-transfection and is maximal 48-72 hours post transfection

CRISPRi lentiviral sgRNA is well suited for long term repression

Reagents & controls

CRISPRi synthetic guide RNA

CRISPRi lentiviral guide RNA

CRISPRi dCas9-SALL1-SDS3

Screening libraries

CRISPRi screening libraries

CRISPRi workflows

Horizon's CRISPRi workflows

CRISPRi workflows