TECHNICAL SUPPORT

Getting Started with CRISPR

Synthego has supported researchers embarking on their first CRISPR experiments for over a decade. Here, we share some of our favorite resources for researchers beginning their CRISPR journeys as well as general information around planning your experiment.

General Resources

What Is CRISPR? Guide: Interested in starting at the beginning? This guide provides a holistic introduction to CRISPR, from its discovery to exciting applications.
CRISPR 101 eBook: Thinking about your first CRISPR experiment? This eBook offers a comprehensive introduction to the molecular biology underlying CRISPR and commonly used CRISPR-based techniques.
CRISPR Knockouts eBook: Want to learn more about CRISPR knockouts (KOs)? This eBook provides a detailed introduction to using CRISPR to inactivate genes.
CRISPR Knock-ins 101 eBook: Looking to learn more about CRISPR knock-ins (KIs)? This eBook offers guidance for creating genomic insertions using CRISPR.
Comprehensive Guide on CRISPR Methods: Curious about CRISPR applications beyond KOs and KIs? This guide discusses other exciting applications for CRISPR technologies, such as prime editing, CRISPRa, and CRISPRi.
CRISPR Delivery Methods: Cargo, Vehicles, and Challenges: Deciding how to deliver CRISPR machinery? This blog discusses the pros and cons of each delivery strategy and introduces new delivery methods on the horizon.
CRISPR Applications: Agriculture, Medicine, Bioenergy, & the Future: Interested in diverse applications for CRISPR? This resource discusses CRISPR as a powerful genome editing tool for medical, agricultural, and other applications.
Alternatives to CRISPR-Cas9: Nucleases for Next-Gen Therapy: Intrigued by CRISPR-Cas nucleases beyond Cas9? This blog discusses nucleases beyond Cas9 with an eye towards their therapeutic use.
History of Genetic Engineering and the Rise of Genome Editing Tools: Wondering how CRISPR fits into the broader picture of genome editing technologies? This resource discusses the history of genome editing, beginning with the discovery of DNA.
Demystifying CRISPR gRNA Chemical Modifications: Curious about chemically-modified synthetic gRNAs? This blog describes common chemical modifications and their benefits for CRISPR experiments.

Planning for Your CRISPR Experiment

Once you are ready to perform your first CRISPR experiment, here are some recommendations for getting started.

Materials

For any CRISPR genome editing experiment, you need the two core components of the CRISPR editing machinery: CRISPR enzyme or Cas nuclease (CRISPR-associated endonuclease) and guide RNA (gRNA).

CRISPR-Cas Systems. Most CRISPR-Cas systems are comprised of a guide RNA (gRNA) and genome editor, which together form a ribonucleoprotein (RNP) complex. Comparative RNP complex schematics of Synthego’s eSpOT-ON, SpCas9, hfCas12Max, and AccuBase are shown here.

CRISPR Enzyme: Synthego offers high-quality genome editors in purified protein and mRNA format. This page introduces each of Synthego’s CRISPR nucleases. Each nuclease has unique qualities, so it is essential to select a nuclease that has the performance characteristics (activity, fidelity, edit type) that you need for your experiments. For therapeutic developers, we also recommend considering licensing early in the development process.
gRNA: Synthego specializes in the production of best-in-class synthetic gRNAs. This page offers a detailed introduction to Synthego’s gRNAs for research applications. To learn more about the benefits of using synthetic gRNAs, we recommend this case study. Your custom synthetic gRNAs will be specifically designed for your experimental goals. For design support, we recommend reviewing our CRISPR gRNA Design Tools page as a starting point.

For experiments using Synthego’s nucleases (eSpOT-ON protein, eSpOT-ON mRNA, hfCas12Max, SpCas9) or cytosine base editor (AccuBase), we offer pre-designed positive control gRNAs. We strongly recommend including these gRNAs in your early experiments to facilitate delivery optimization.

Methods

Delivery: Your CRISPR machinery (gRNA and nuclease) must enter the target cells to execute their genome-editing functions. Synthego’s CRISPR solutions can be delivered into your target cell population using many transfection methods. Protocols for popular transfection methods are below.
- Nucleofection: SpCas9, hfCas12Max, eSpOT-ON (nuclease protein, nuclease mRNA)
- Neon Electroporation: SpCas9
- Lipofection: SpCas9
- A full list of protocols may be found on our Resources page.
The optimal transfection conditions are experiment-dependent. Please view these protocols as a starting point. Optimization will be required to obtain the best possible editing efficiency in your system.
Analysis: To determine the success of your CRISPR experiment, we recommend using a two-pronged strategy to assess your genomic edits.
1. Genotyping: This will allow you to determine the edits present in the genomic DNA of your target cells. We recommend using a sequencing-based method, specifically next-generation sequencing or Sanger sequencing plus ICE analysis. For more information about ICE, we recommend our Genotyping with ICE Protocol and our Step-by-Step Guide for Analyzing CRISPR Editing Results with ICE.
2. Phenotypic or functional assay This will allow you to confirm that your edit has the desired effect. The phenotypic or functional assay best for your experiment depends on your CRISPR edit and experimental system. For knockout experiments, our Protein Analysis of Knockouts guide includes an introduction to commonly used assays.

Additional Assistance

Having trouble with your gRNA designs? Connect with our scientific support team!

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