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.
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.
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.
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.
- 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.
- Transform the CRISPR vector into competent E. coli cells (available in kits) to generate sufficient expression plasmid.
- 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.
Order GeneArt CRISPR Nuclease Vector Kits for designing CRISPR plasmids
Discover how your CRISPR gene editing experiments can benefit from Cas9 proteins and how the popular nuclease compares to Cas9 plasmids.
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
For Research Use Only. Not for use in diagnostic procedures.