All-in-one CRISPR plasmid system to transfect and select edited cells

For designing all-in-one CRISPR-Cas9 vectors, use Invitrogen GeneArt CRISPR Nuclease Vector Kits, which incorporate both a Cas9 nuclease cassette and a targeted guide RNA (gRNA) cloning cassette, allowing you to edit a genomic locus of choice from a single Cas9 plasmid.

The kits include tools for CRISPR-Cas9 vector design, construction, and propagation, along with reporters that enable sorting or enrichment of transfected populations.

Order the GeneArt CRISPR Nuclease Kit

Combining Cas9 nuclease, gRNA, and reporters in a single CRISPR-Cas9 vector


GeneArt CRISPR Nuclease Vector Kits make it easy to express a guide RNA (including crRNA and tracrRNA) using a plasmid vector that also expresses Cas9 endonuclease. Choose the GeneArt CRISPR Nuclease Vector with OFP (orange fluorescent protein) for flow cytometry-based sorting of crRNA-expressing cell populations, or the GeneArt CRISPR Nuclease Vector with CD4 for bead-based enrichment of crRNA-expressing cells.
 

Image of product box and tubes for the GeneArt CRISPR Nuclease Vector with Orange Fluorescent Protein Reporter Kit

TIP: If you’re new to CRISPR editing, we recommend using TrueCut Cas9 Protein with a TrueGuide Synthetic gRNA rather than a Cas9 DNA plasmid. The ribonuceoprotein complex becomes active immediately upon transfection and is cleared from the nucleus faster than the CRISPR plasmid, reducing the opportunity for off-target effects or DNA integration.

Plasmid maps of Cas9 nuclease expression vectors with OFP reporter or CD4 reporter.

Figure 1. GeneArt CRISPR nuclease vector maps. The vector is supplied prelinearized with 5-bp 5’ overhangs as shown for easy cloning of your double-stranded DNA oligo that encodes a target-specific crRNA. Maps are shown of the vectors with (A) OFP reporter and (B) CD4 reporter. The gRNA, Cas9, and reporter are expressed from the same vector. Cas9 is directed to the nucleus by nuclear localization signals (NLS1 and NLS2).
 

Creation and expression of CRISPR plasmids using the GeneArt CRISPR Nuclease Vector Kit

Figure 2 summarizes the three-step workflow to create a CRISPR vector and express it in cells. 

 

  1. Design single-stranded DNA oligonucleotides that code for the target-specific portion of your CRISPR gRNA. Anneal and clone these DNA oligos into the CRISPR-Cas9 vector. Use the TrueDesign Genome Editor software to determine an optimal gRNA target region for your desired edit.




  2. Transform the CRISPR vector into competent E. coli cells (available in kits) to generate sufficient expression plasmid.




  3. Transfect the target cells with the CRISPR vector and use the OFP or CD4 reporter to sort or enrich the transfected cells for further use.
     

Figure 2. Workflow for utilization of GeneArt CRISPR Nuclease Vector Kit. Following annealing and cloning of DNA oligos coding for target-specific crRNA into a Cas9 nuclease reporter vector, the vector is transformed into E. coli cells. E. coli are screened for the desired CRISPR clone, which is then transfected into your cells for gene editing and analysis.
 

  Download the GeneArt CRISPR Nuclease Vector Kit user guide

Order GeneArt CRISPR Nuclease Vector Kits for designing CRISPR plasmids

INFOGRAPHIC: Cas9 plasmids vs proteins

Discover how your CRISPR gene editing experiments can benefit from Cas9 proteins and how the popular nuclease compares to Cas9 plasmids.

Download infographic

Products for creating your own CRISPR vectors

In addition to using our GeneArt CRISPR Nuclease Vector Kits for an all-in-one CRISPR plasmid system, you may choose to use separate Cas9 plasmids and gRNA plasmids.

Plasmids for Cas nuclease expression and gRNA cloning vectors can be obtained from open-access resources (e.g., Addgene, the nonprofit global plasmid repository). For constructing your own plasmid for gRNA expression, you may need oligonucleotides containing your gRNA sequence, as well as FastDigest restriction and cloning enzymes.
 

Additional products for your CRISPR plasmid workflow

Products for transformation of CRISPR plasmids

We offer a variety of competent E. coli cells for propagation of your Cas9 plasmid, including One Shot MAX Efficiency DH5a T1R Chemically Competent Cells as well as the PureLink Expi Endotoxin-Free Maxi Plasmid Purification Kit for plasmid isolation and purification.
 

Products for CRISPR plasmid delivery into cells and validation of CRISPR edits

Transfection products for effective delivery of CRISPR vectors
Methods to validate your gene editing results
Cas9 Proteins 
 

Resources and support for CRISPR plasmids

Molecular Cloning Education
Cloning Support Center
 

Gene editing resources and support
Resources
Support

For Research Use Only. Not for use in diagnostic procedures.