Question: Which 10x Genomics Single Cell CRISPR approach should I use?
Answer: 10x Genomics single cell products offer a variety of ways to measure gene expression while also identifying CRISPR perturbations through direct detection of guide RNAs.
- Our single cell 5’ product allows direct detection of guide RNAs through targeted reverse transcription of the sgRNA molecules themselves.
- Our Flex assay uses custom probes to report the presence of guides.
Which approach is best for your experiment depends on a few factors.
Background
CRISPR screens are the preeminent way for performing functional genomics. Using RNA-guided endonucleases (or modified versions with other engineered activities including silencing and activation) researchers can perform targeted genetic perturbations at massive scale in pooled format.
While traditionally CRISPR screens have been performed with some form of selection as a readout (growth, death, sensitivity or resistance to a particular treatment, expression of a reporter) more recently, researchers have begun to perform high throughput CRISPR screens with more detailed single cell molecular phenotypes as readouts.
While some methods (especially those involving combinatorial barcoding) are limited to measuring perturbations by the presence of large lowly expressed transcripts from specially engineered constructs (e.g. CROP-seq style detection), methods that directly detect the small but very highly expressed guide RNAs themselves afford vastly higher guide recovery and much more robust calling of perturbations, leading to better data and greatly enhanced overall experimental efficiency.
GEM-X Universal 5’ Gene Expression with Feature Barcode Technology for CRISPR Screening
For most customers, our 5’ assay is the best fit as you are beginning your single cell functional genomics assays. With overloading afforded directly by the nature of pooled CRISPR screens, researchers can achieve costs of ~$0.025 USD per single cell† and ~45,000 cells per lane (or 360,000 cells per chip) with standard workflows that produce high complexity gene expression and guide RNA libraries from the same single cells.
This scale easily accommodates customers who want to perform screens with hundreds to low thousands of perturbations with low hundreds of cells observed per perturbation.
GEM-X Flex Gene Expression
For some customers, our Flex product with custom probes directed against guide RNAs may be the appropriate choice. Our newly released n-Plex kit allows customers to profile up to 5M cells at <$0.01 USD per single cell†. Pairing this with probe-mediated guide detection provides cost and scale unimaginable with other approaches at a quality that has independently been determined to be clearly market leading in both quality and sequencing efficiency.
Use cases where it makes sense to consider Flex with custom probes include:
- Screens that require cell numbers into the millions of cells and up including when customers want to
- Screen 10s of thousands of perturbations
- Have high numbers of observations of each perturbation (such as when the expected phenotypes are more subtle)
- Repeated screening of the same perturbations across a number of conditions, cell types, timepoints
- Screens where fixation may be required or desirable
Please refer to our knowledge base articles for support on designs and necessary workflow and analysis adjustments for Flex+CRISPR:
- What workflow modifications are required when performing CRISPR Screening with the Single Cell Gene Expression Flex assay?
- How do I design custom probes for CRISPR Guide Capture for use with the Single Cell Gene Expression Flex Assay?
- How do I pool custom probes for a Single Cell Gene Expression Flex + CRISPR experiment?
- How do I analyze CRISPR screening + Single Cell Gene Expression Flex data in Cell Ranger?
† Cost figures represent 2024 US list prices. For further cost breakdown please reach out to your local sales representative
Product: Single Cell Immune Profiling, Single Cell Gene Expression Flex