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Sanger Whole Genome CRISPR Arrayed Libraries

Designed by the Wellcome Trust Sanger Institute




  Sanger Arrayed Lentiviral CRISPR Libraries
  won prestigious R&D 100 Award

 The Power of Gene Knockout!

Two leaders in genome editing, MilliporeSigma and The Wellcome Sanger Institute, have joined forces to make the first ever arrayed lentiviral CRISPR knockout libraries for human and mouse genomes.

Genome-wide loss-of-function screening is a powerful approach to discover genes and pathways that underlie biological processes. Now complete knockout is achievable with two optimized gRNAs per gene. Minimized clone number ensures the most specific screening possible while controlling time and cost.

Early screening experiments used RNA interference (RNAi), which enables gene knockdown but not complete gene knockout. Thus, RNAi may miss important gene hits when total loss of gene expression is required. CRISPR-Cas9 technology allows more efficient loss-of-function screening in mammalian cells on a large-scale.

The advent of pooled CRISPR libraries brought efficient genome-wide knockout but required the additional step of deep sequencing deconvolution for accurate hit identification. Arrayed screening has the advantage of easy hit identification with no deconvolution necessary. Arrayed CRISPR screening also enables high content assays such as cellular imaging, fluorescence, luminescence and colorimetric experiments. Adapting the CRISPR-Cas9 technology, genome-wide, the exclusive MilliporeSigma Sanger CRISPR libraries greatly expand high throughput screening capabilities.
 

 The Next Generation of Screening Tools has Arrived

Achieve efficient, long term knockout, even in difficult-to-transfect cell lines.

The CRISPR-Cas9 system for genome editing greatly expands research capabilities. Now discover and characterize gene function with rapid screening and analysis of thousands of gene editing events in a single experiment.

Using a validated approach tested at Sanger and published in the literature, the Sanger arrayed CRISPR Libraries are cutting-edge technology, now available to all. Perform initial pilot experiments and optimize your cellular system and assays with our CRISPR controls. Exclusive to MilliporeSigma, the Sanger arrayed libraries are a unique complement to our existing CRISPRRNAi and ORF libraries. With this addition, we continue to provide the most advanced CRISPR portfolio for your screening experiments, from knockdown and knockout to expression and validation.

Visit Nature Biotechnology to read more on the use of RNAi and CRISPR to build a complete research story.

 Start Your CRISPR Screen Today

Why troubleshoot false positive results when you can use the gRNA-optimized, arrayed Sanger CRISPR Library? The Sanger CRISPR library provides extensive, high-quality gene knockout and is available only at MilliporeSigma. Fast-track your research with the first whole genome arrayed library available!

Optimal Lentiviral-Based CRISPR

  • Stable or transient transfection
  • Enrich population of cells (select puromycin- and BFP-positive hits)
  • Viral particles for sustained integration, creating a stable cell line
  • Transduces virtually any cell type (dividing or non-dividing cell lines)
  • Custom titers available for any application
  • BFP tagged vector for positive-cell visualization
Figure 1. BFP expression in A549-Cas9-Blast
stable cells infected with arrayed CRISPR
lenti-gRNA viruses, seven days post infection,
20X magnification.

Need help with CRISPR fundamentals? Learn more about Getting Started with CRISPR.

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 Sanger Library Specs

Glycerol Format

  • Approximately 400 96-well plates, 50 µl bacterial glycerol stock per clone
  • Human or mouse species

Lentiviral Format

  • Approximately 100 384-well barcoded labeled Costar plates with pierceable aluminum seals
  • 10 ul per clone at a minimum titer of 1x106 particles/ml as determined by p24 assay
    Human or mouse species

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 Sanger CRISPR gRNA Design and Vector Features

gRNA Design

  • Maximizes gene knockout by targeting the first half of the protein coding sequence
  • Optimizes targeting of all transcripts
  • Avoids the first 90 bp of coding sequence where alternative start codons often reside
  • Sequences are highly conserved, avoiding SNPs, to ensure representation in multiple cell lines
  • Stringent rules reduce or eliminate the potential for off-target effects

Vector Features

  • Simplify the workflow with puromycin selection
  • Illuminate CRISPR-expressing cells and enrich the transduced population with BFP
  • Transposase sites provide an alternative to lentiviral delivery

Figure 2. Sanger Lentiviral CRISPR vector schematic (U6-gRNA:PGK-puro-2A-tagBFP)

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 Lentiviral Production Expertise

MilliporeSigma is committed to providing unmatched manufacturing and support for genome editing technology. With state-of-the-art robotics, liquid handling, LIMS and dedicated laboratories in our Life Science and High Technology Center, we provide superior quality and convenient service for your CRISPR research needs.

High Quality Manufacturing Systems and Personnel

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 Screen at Any Scale

The Sanger library is the only tool that allows both whole genome CRISPR KO interrogation and sophisticated phenotypic readout in a convenient arrayed format. For smaller scale experiments, leverage the design expertise of the Wellcome Trust Sanger Institute and manufacturing quality of MilliporeSigma to build a custom panel for your research.

Do you need more focused or affordable screens?

  • Quick pick a single Sanger clone for any gene
  • Cherry-pick a custom Sanger panel of any size
  • Target any pathway or gene family with a Sanger panel
  • Order any gRNA design in the Sanger vector
  • Apoptosis
  • Cancer Biology
  • Cell Cycle
  • Cytokine Receptors
  • Cytokines
  • Developmental signaling pathways
  • DNA Damage Repair
  • Drug Transport
  • Druggable Genome
  • Epigenetic regulators
  • GPCRs
  • Helicase
  • Ion Channels
  • JAK-STAT
  • Kinases
  • Membrane Trafficking
  • Nuclear Hormone Receptors
  • p53
  • Phosphatases
  • Proteases
  • T-Cell Activation
  • Transcription Factors
  • Tumor Suppressors
  • Tyrosine Kinase
  • Ubiquitin Hydrolases
  • Ubiquitin Ligases
  • Whole Genome

Don’t see the option you need? Specify your own gRNA for manufacturing in the Sanger backbone. From whole genome libraries to individual clones, the Sanger CRISPR lentiviral collection is your path to unlimited discovery.
Email crispr@sial.com for more information.

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 Sanger gRNA Application Data

 
A.
  Non-infected cells Infected with virus

 

B.

 

C. gRNA Activity
by CEL-1 %
    Individual gRNA Design Ranked by Cel-1 activity

Figure 4. The CEL-I assay. Sanger lenti-gRNA library demonstrates definitive cleavage. A549-Cas9-Blast cells were infected with a representative 96-well Sanger plate, selected with puromycin and CEL-I assay performed. A) Cell growth after puromycin selection for four days; B) Representative gel from CEL-I Assay; C) High average CRISPR nuclease activity and high frequency of active gRNA clones genome-wide. Each dot represents a single clone.

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References

  1. R. Barrangou et al., Science 315, 1709–1712 (2007).
  2. L. Cong et al., Science 339, 819–823 (2013).
  3. P. Mali et al., Science 339, 823–826 (2013).
  4. W. Y. Hwang et al., Nat. Biotechnol. 31, 227–229 (2013).