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Alt-R™ CRISPR gRNA Libraries
For researchers looking for better flexibility and higher performance CRISPR screening libraries, IDT’s custom synthetic gRNA libraries offer a highly customizable option, strong CRISPR expertise and support, and an optimized RNA synthesis process.
Custom libraries for better CRISPR screening.
Overview
Alt-R Custom CRISPR gRNA Libraries
Custom arrayed synthetic guide RNA libraries are a great way to accomplish CRISPR knockout screens. They accelerate your research by avoiding the cloning and sequencing associated with lentiviral screens while also integrating into existing automation workflows. IDT has experience designing and manufacturing arrayed panels of gRNAs in a range of sizes, from a few guides to thousands or more. Our high-capacity manufacturing systems offers delivery of your custom CRISPR panel starting at as fast as five business days.
- Highly customizable CRISPR libraries: Different gRNA formats (2-part or sgRNA), plate type, custom formulations, and lab automation platforms, to suit a variety of project needs
- Reliable, consistent solution offered by a global leader in RNA synthesis and innovation
- Adaptable for alternative CRISPR systems such as Cas12a, Cas13, and prime editing
- Enhanced nuclease resistance for maximal editing in Cas nuclease-expressing cells or via ribonucleoprotein (RNP) delivery
We know your time is valuable and we’ll prioritize your inquiry. Click on “Request consultation” to provide brief information about your CRISPR library project or interests, and we’ll be in touch to discuss your project ASAP.
Product details
Alt-R Custom CRISPR gRNA libraries are available for all CRISPR nucleases, including Cas9, Cas12a, Cas13, prime editing enzymes, and other alternative systems. These libraries were developed to address the need for better CRISPR screening solutions. They are chemically modified guide RNAs synthesized on IDT’s proprietary, high-fidelity RNA manufacturing platform to provide high quality, reliable gRNA libraries with fast delivery.
| Features | Options |
|---|---|
| Design | Predesigned, custom, user-provided |
| CRISPR systems | Cas9, Cas12a, Cas13, prime editing, and other alternative systems |
| Guaranteed yield | 0.5 nmol, 2 nmol, 5 nmol, and custom normalized deliverables |
| Cas9 gRNA formats | 2-part crRNA:tracrRNA complex and sgRNA |
| Custom lengths supported | 30–150 nt |
| Chemical modifications | 2’-O-methyl RNA, PS linkages, end-blocking Alt-R modifications |
| Plate types | 96- & 384-well PCR, Deep-well, V-bottom, Echo, custom options available |
| Formulation options | Multi-guide per well; pooled by gene Arrayed (single gRNA/well) Custom formulations upon request |
| QC | Individual ESI/MS |
| Supporting reagents & functional analysis pipeline (optional) | WT Cas9, HiFi Cas9, Cas12a. and Cas12a Ultra Glycerol-free options available in tubes or plates (ideal for robotics) Electroporation Enhancers rhAmpSeq™ CRISPR Analysis System (NGS-based on-/off-target editing analysis) |
Alt-R CRISPR Enzymes
Our Alt-R CRISPR enzymes are available in a variety of formats, with stock sizing available up to 50 mg. Larger formats and lot matching are also available for all products upon request.
| Cas protein | Available versions | Key features |
|---|---|---|
| S.p. Cas9 Nuclease V3 | Wild-type, 50% Glycerol Wild-type, Glycerol-free High Fidelity (HiFi) Wild-type Fluorescent-fusion | Targeting GC-rich regions Low viscosity for high-throughput applications Reduced off-target activity* Fluorescent label for enrichment (GFP or RFP) |
| A.s. Cas12a (Cpf1) V3 | Wild-type Ultra | Targeting AT-rich regions Increased on-target activity* |
| L.b. Cas12a (Cpf1) | Ultra | Increased on-target activity and low-temperature tolerance* |
* when compared to corresponding Wild-type controls
Product data
Alt-R guide RNA libraries and CRISPR products offer an efficient solution for creating knockout libraries
We investigated the efficiency of inducing non-homologous DNA end joining (NHEJ) by designing 3 sgRNAs per gene (N=95 genes) via an internal pipeline and using IDT’s Alt-R Cas9 nuclease S.p. Cas9 WT V3, electroporation enhancer, and rhAmpSeq™ CRISPR analysis system. The combined use of the Alt-R guides and additional CRISPR products provided an effective, high-throughput editing solution with >90% NHEJ editing in 99% of targeted genes (Figure 1).
Figure 1. A 3-guide-per-gene library design resulted in >90% NHEJ editing in almost all (90/91) targeted amplicons.
95 genes of interest were fed into an internal design pipeline to generate 3 guide RNA designs per gene, all
located within a 500 base span. Once target sites were identified, singleplex genotyping assays for each gene were selected using the rhAmpSeq Design Tool. K562 cells were transfected with 3 sgRNAs complexed to Alt-R S.p. Cas9 WT Nuclease V3 per well
(final concentration = 1 µM per guide). rhAmpSeq CRISPR Library preparation and subsequent sequencing on the Illumina MiSeq platform was then performed. IDT’s rhAmpSeq Analysis Tool revealed a high level of NHEJ-type editing events (dark
blue) in nearly all amplified regions (N = 91 amplicons with sequencing reads >500).
Alt-R CRISPR-Cas9 sgRNAs provide effective editing in Jurkat cells
To highlight the editing efficiency of sgRNAs, we designed sgRNAs targeting 255 sites across the human genome and delivered them to Jurkat cells (a human T-lymphocyte-derived cancer cell line) along with Alt-R S.p. WT Cas9 Nuclease V3. The result of this experiment shows that Alt-R CRISPR-Cas9 sgRNAs provide high levels of editing.
Figure 2. High levels of editing with Alt-R CRISPR-Cas9 sgRNAs.
Ribonucleoprotein (RNP) complexes were formed with Alt-R S.p. WT Cas9 Nuclease V3, combined with Alt-R Cas9 sgRNAs synthesized for 255 randomly selected Cas9 guide RNA sites across the human genome. RNP complexes (4 μM) were delivered into Jurkat cells via a Nucleofector™ system (Lonza) in the presence of Alt-R Electroporation Enhancer. Genome editing efficiencies were determined by target amplification followed by next generation sequencing (NGS) on an Illumina™ instrument.
Resources
Frequently asked questions
Does IDT offer CRISPR screening analysis solutions?
Yes, we have integrated analysis options.
Please find our advanced data analysis pipeline for quantifying your genome editing results here: rhAmpSeq™ CRISPR Analysis System
Please find our advanced data analysis pipeline for quantifying your genome editing results here: rhAmpSeq™ CRISPR Analysis System
Does IDT offer CRISPR library design services?
- IDT's online CRISPR-Cas9 gRNA design tools can be used to assist with designs for Cas9 libraries. Please find our design tool to generate custom CRISPR-Cas9 guide RNAs here: Custom Alt-R™ CRISPR-Cas9 gRNA Design Tool
- For complex designs please contact us.
I want to target each of my genes with multiple guides. How many gRNAs do you suggest per gene?
While we currently do not have data to support a definitive answer, our R&D team routinely use 2-3 guides per gene for Cas9 editing projects with high editing success rates. Likewise, most customers typically prefer 2-4 guides per gene, depending on the project.
What synthetic arrayed library formats are available from IDT? Are additional customization options possible?
- Table 1 shows available customizations for library design, formulations, and shipping formats.
Product Specifications
| Features | Options |
|---|---|
| Design | Predesigned, custom, user-provided |
| CRISPR systems | Cas9, Cas12a, Cas13, prime editing, and other alternative systems |
| Guaranteed Yield | 0.5 nmol, 2 nmol, 5 nmol, and custom normalized deliverables |
| Cas9 gRNA formats | 2-part cRNA:tracrRNA complex and sgRNA |
| Custom lengths supported | 30-150 nt |
| Chemical modifications | 2'-O-methyl RNA, PS linkages, end-blocking Alt-R modifications |
| Plate types | 96- & 384-well PCR, Deep-well, V-bottom, ECHO, custom options available |
| Formulation options | Multi-guide per well; pooled by gene Arrayed (single gRNA/well) Custom formulations upon request |
| QC | Individual ESI/MS |
| Supporting reagents & functional analysis pipeline (optional) | WT Cas9, HiFi Cas9, Cas12a, and Cas12a Ultra Glycerol-free options available in tubes or plates (ideal for robotics) Electroporation Enhancers rhAmpSeq™ CRISPR Analysis System (NGS-based on-/off-target editing analysis) |
- Don’t see what you’re looking for? We are continually expanding our CRISPR library products and we may have what you need. If you are interested in other chemically modified gRNAs (such as CRISPR on/off systems) targeting any sequence from any species, email our CRISPR experts today to discuss customized solutions for your research at CRISPR@idtdna.com.
What are the advantages of an arrayed CRISPR screen over a pooled screen?
- Unlike pooled libraries, knowing exactly which gene was targeted in a population of cells negates the need for a sequencing-based deconvolution step. Additionally, arrayed libraries are amenable to phenotypic-based high-throughput screens because there are no confounding effects from multiple gene knockouts.
- Synthetic arrayed gRNA libraries can be delivered directly to cells without viral packaging, avoiding any biosafety issues posed by using lentiviruses.
What is the difference between arrayed and pooled CRISPR gRNA libraries?
- Both types of libraries consist of guides targeting a wide range of genes or other loci. There can be a single guide per gene, or multiple.
- Pooled library guides are all mixed together and delivered to a bulk population of cells, often via lentiviral transduction.
- Arrayed libraries deliver guides targeting one gene to an individual population of cells. For example, you can have 96 distinct cell populations, each with a different gene knock-out.
What type of CRISPR guide RNA libraries does IDT offer?
- IDT provides arrayed synthetic gRNA in many different formats and library projects can range from 10s to 100s to 1000s of guides.
- A synthetic guide means it is already an RNA, so no need to perform in vitro transcription or clone into an expression plasmid.
- If you're interested in pooled lentiviral libraries or performing transcription yourself, IDT can also provide single-stranded DNA oPools™ Oligo Pools for cloning.
Do you offer CRISPR crRNA libraries?
Yes, we offer predesigned Cas9 CRISPR crRNA and additional options for creating customizable gRNA libraries.
For more information, please visit our Custom guide RNA Libraries page.
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