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All-in-one Cas9 SmartNuclease™ & AAVS1 gRNA Plasmid

Streamline genome editing at the AAVS1 Safe Harbor Site with a plasmid that expresses both Cas9 and a gRNA that targets the AAVS1 site

  • Easy, precise knock-in of any gene
  • Consistent, robust transgene expression from the AAVS1 Safe Harbor Site
  • Simplified construction of isogenic cell lines
  • Minimal off-target integration when using our AAVS1 HR Targeting Vectors
  • Streamlined CRISPR/Cas9 library screening
Catalog Number
Description
Size
Price
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Add to Cart
CAS601A-1
All-in-one plasmid to express Cas9 SmartNuclease and gRNA targeting the AAVS1 Safe Harbor Site (EF1α-hspCas9-H1-AAVS1_gRNA)
10 µg
$ 936

Overview

Streamline genome editing at the powerful AAVS1 Safe Harbor Site

When you have transfectable cells and want to take advantage of the robust and reliable expression that’s possible from the AAVS1 Safe Harbor Site, SBI offers a streamlined plasmid system that’s ready to go. The All-in-one Cas9 SmartNuclease™ & AAVS1 gRNA Plasmid expresses both Cas9 and an AAVS1-targeting gRNA—just co-transfect with a homologous recombination (HR) Targeting Vector and select for your desired colonies.

Delivering consistent, robust transgene expression, the AAVS1 safe harbor site is a preferred target for gene knock-ins. Insertion at the site has been shown to be safe with no phenotypic effects reported, and the surrounding DNA appears to be kept in an open confirmation, enabling stable expression of a variety of transgenes.

SBI’s AAVS1 Safe Harbor Targeting products deliver:

  • Easy, precise knock-in of any gene
  • Consistent, robust transgene expression from the AAVS1 Safe Harbor Site
  • Simplified construction of isogenic cell lines
  • Minimal off-target integration when using our AAVS1 HR Targeting Vectors
  • Streamlined CRISPR/Cas9 library screening

Note that we’ve specially designed a series of AAVS1 HR Targeting Vectors that greatly reduce off-target integration events (Cat.# GE620A-1, GE622A-1, and GE624A-1). Taking advantage of the AAVS1’s location within an intron, these vectors come with a puromycin marker that has no promoter, only a splice acceptor site—expression of puromycin can only occur when the construct integrates within an intron, reducing the probability of recovering off-target integrants in the presence of puromycin selection.

As with all of our Cas9 delivery options, the All-in-one Cas9 SmartNuclease & AAVS1 gRNA Plasmid is functionally validated and comes backed by our expert technical support team—if you’ve got a genome engineering question just ask by emailing tech@systembio.com.

Not sure whether you need a CRISPR/Cas9 plasmid, purified protein, or mRNA?

Use this table to choose the CRISPR/Cas9 product that’s right for you:

For This Application
In these types of cells

Use These Products

MODIFYING ORGANISMS

  • Gene tagging
  • Transgenic organism generation
  • Model organism engineering
Embryos—to create transgenic animals Injectable Cas9 mRNA & gRNA Synthesis Kits
Cas9 Protein
EGFP-labeled Cas9 Protein
Animals models—in vivo genome editing AAV-Cas9 Vectors  
Cas9 Protein
EGFP-labeled Cas9 Protein
MODIFYING CELL LINES

  • Stable KO, KI, and genome editing of
    somatic cells
  • Transgenic cell line generation
  • Cell-based disease models
Cells that are transfectable Cas9 Plasmids
Cas9 Protein
EGFP-labeled Cas9 Protein
Difficult-to-transfect cell lines:

  • Primary cells
  • Hematopoietic cells
  • Stem cells
AAV-Cas9 Vectors 
Lenti Cas9 Systems
SCREENING

  • Genome-wide surveys
  • gRNA library screens
  • Functional screens
All cell types requiring stable Cas9 overexpression Lenti Cas9 Systems
AAVS1 Safe Harbor Site Cas9
Gene Knock-in System
Cas9 Protein
EGFP-labeled Cas9 Protein
PRE-CLINICAL APPLICATIONS

  • Off-target events are of highest concern
All cell types and applications Cas9 Nickase, available in all delivery formats
Cas9 Protein
EGFP-labeled Cas9 Protein
SIMULTANEOUS ENGINEERING OF MULTIPLE MUTATIONS All cell types and applications Multiplex gRNA cloning kit, compatible with all Cas9 delivery options

How It Works

Genome engineering with CRISPR/Cas9

For general guidance on using CRISPR/Cas9 technology for genome engineering, take a look at our CRISPR/Cas9 tutorials as well as the following application notes:

CRISPR/Cas9 Gene Knock-Out Application Note (PDF) »
CRISPR/Cas9 Gene Editing Application Note (PDF) »
CRISPR/Cas9 Gene Tagging Application Note (PDF) »

CRISPR/Cas9 Basics

Through careful selection of the target sequence and design of a donor plasmid for homologous
recombination, you can achieve efficient and highly targeted genomic modification with CRISPR/Cas9.

The system

Cas9 protein—uses guide RNA (gRNA) to direct site-specific, double-strand DNA cleavage adjacent to a protospacer adapter motif (PAM) in the target DNA.

gRNA—RNA sequence that guides Cas9 to cleave a homologous region in the target genome. Efficient cleavage only where the gRNA homology is adjacent to a PAM.

PAM—protospacer adapter motif, NGG, is a target DNA sequence that spCas9 will cut upstream from if directed to by the gRNA.

The workflow at-a-glance

DESIGN: Select gRNA and HR donor plasmids. Choice of gRNA site and design of donor
plasmid determines whether the homologous recombination event results in a knock-out,
knock-in, edit, or tagging.

CONSTRUCT: Clone gRNA into all-in-one Cas9 vector. Clone 5’ and 3’ homology arms into HR
donor plasmid. If creating a knock-in, clone desired gene into HR donor.

CO-TRANSFECT or CO-INJECT: Introduce Cas9, gRNA, and HR Donors into the target cells
using co-transfection for plasmids, co-transduction for lentivirus, or co-injection for mRNAs.

SELECT/SCREEN: Select or screen for mutants and verify.

VALIDATE: Genotype or sequence putative mutants to verify single or biallelic conversion.

Citations

  • Ihry, RJ, et al. (2018) p53 inhibits CRISPR-Cas9 engineering in human pluripotent stem cells. Nat. Med.. 2018 Jun 11;. PM ID: 29892062

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