CRISPR Gene Editing Workflow: Confirmation of Edits

Solutions for Confirmation of Gene Edits

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CRISPR gene editing provides versatile workflows that enable researchers to edit their desired target genes efficiently and accelerate therapeutic development. Explore our solutions for gene editing workflows.

Confirming Targeted Gene Edits

After cells of interest have been enriched, it is important to confirm that the target cells have been successfully edited prior to moving on to downstream assays. This can be accomplished either through direct detection of edits using genomic methods or through indirect detection using cellular or proteomic methods.

PCR-based methods provide precise and sensitive detection and quantification
of edits. High-fidelity PCR, Droplet Digital PCR (ddPCR), real-time PCR, and high
resolution melt analysis (HRMA) allow analysis of NHEJ and HDR edits. The CFX
Real-Time PCR Detection System is a versatile platform for quantitative PCR and HRMA. For standard PCR and sequencing validation, iProof High-Fidelity PCR Reagents minimize the incorporation of errors. When editing efficiencies are low, ddPCR technology allows identification of allele changes with unmatched precision; mutations present at frequencies ≤0.5% can be identified.

The ZOE Fluorescent Cell Imager allows confirmation of knocked in and knocked out protein expression in newly edited cells right at your bench. The V3 Western Workflow quantitatively measures protein expression faster and with better accuracy than other western blot procedures. By incorporating validated PrecisionAb Western Blotting Antibodies, uncertainty can be removed from experiments.

The ZOE Cell Imager and V3 Western Workflow can be used to confirm successful gene edits. These orthogonal methods, along with direct methods such as Droplet Digital PCR or HRMA using qPCR provide the confidence to move forward with downstream analysis of edited cells.

Companion Products for Gene Editing Confirmation

  • ddpcr-prod-nav-systems

    QX200 Droplet Digital PCR System

    • Detection of small or large edits with absolute quantification
    • Most precise and sensitive digital PCR solution for all applications
    • Flexible digital PCR chemistry — optimized for TaqMan Hydrolysis Probe and EvaGreen Assays
    • Flexible assay setup for high sensitivity or high throughput
  • qPCR

    CFX Real-Time PCR Detection System

    • Sensitive, reliable detection for singleplex and multiplex real-time PCR reactions
    • 2–5 color multiplexing, advanced optical technology, and precise gradient temperature control
    • CFX Maestro Software for superior data collection, visualization, and statistical analysis
  • pcr-master-mixes-kits-thumb-standard-pcr-reagents

    iProof High-Fidelity Real-Time PCR Reagents

    • High fidelity — iProof DNA polymerase is 52-fold more accurate than Taq polymerase
    • Speed — high processivity dramatically reduces extension (15–30 sec/kb) and overall run times
    • Successful amplification of long products with higher yields — fragments up to 37 kb are amplified in less time and with less enzyme (0.25–1 unit/reaction)
  • Real-Time_PCR Supermixes_and Kits

    Real-Time PCR Supermixes and Kits

    • Any instrument — universal reference dye is compatible with all qPCR platforms
    • Any chemistry — supermixes for SYBR Green or probe-based detection chemistry
    • Any conditions — our patented Sso7d fusion polymerase guarantees superior qPCR performance under varying and/or challenging conditions
  • V3 Western Workflow

    V3 Western Workflow — Visualize, Verify, Validate

    A five-step streamlined western blotting protocol with innovative tools including proprietary stain-free imaging:

    • Faster run and transfer times
    • Checkpoints at every step
    • Quantitation made simple
  • PrecisionAb Validated_Western_Blotting_Antibodies

    PrecisionAb Validated Western Blotting Antibodies

    • Robust validation — performed on up to 12 different cell lysates with endogenous protein levels
    • Reproducible data — stringent batch-to-batch QC
    • High specificity and sensitivity — low nonspecific binding and strong signal from target proteins
  • qPCR

    ZOE Fluorescent Cell Imager

    • Simplified cell imaging — the intuitive touch-screen interface allows users to view cells, capture images, and create multichannel merges with minimal training
    • Flexible operation — brightfield and three fluorescence channels enable use for routine cell culture applications and more sophisticated imaging applications