Transgene Knockin via CRISPR at AAVS1 and ROSA26 Loci
Transgene expression via random integration into the genome is subjected to position effects and silencing. In addition, random gene insertion might interrupt or activate the neighboring genes. Genomic safe harbor sites are transcriptionally active, therefore allowing robust and stable gene expression. Furthermore, a transgene insertion at genomic safe harbors does not have adverse effect on the host cell genome. CRISPR technology can be utilized to do targeted gene insertion at these genomic loci. For Human cells, AAVS1 has been accepted as a high gene expression and a safe genomic location; for Mouse cells, ROSA26 is proved to be a genomic safe harbor locus.
Why is Safe-harbor Transgene Insertion Needed?
- Targeted transgene insertion
- Robust and stable gene expression
- No random genetic interruption and insertional mutagenesis
- Critical for therapeutic cell engineering and many functional screening/assays
- Genome-wide TrueORF collection serves as the source for transgenes
- Easy shuttling from TrueORF clones into AAVS1 and Rosa26 donor vector ( Precision Shuttling System)
gRNA/CRISPR cuts the genome, then the gene of interest (GOI) cassette will be knocked in via homologous recombination.
AAVS1 Safe Harbor knockin system via CRISPR
This system needs two components:
- AAVS1 gRNA vector,All-in-one CRISPR vector containing Cas9 expression cassette and specific gRNA targeting AAVS1 locus
- AAVS1 Donor vector,donor vector for transgene integration, containing the AAVS1 homologous arms (Gene of interest needs to be cloned)
AAVS1 gRNA Vector
OriGene offers a prevalidated AAVS1 gRNA and many AAVS1 donor vectors, which contains different combination of drug selection marker and promoter. In addition to the individual gRNA and donor vector, the pre-assembled AAVS1 vector kit is also available, including not only AAVS1 specific gRNA and donor vector, but also a non-specific scramble control. You can also get our predesigned positive donor control for your experiment or request a custom AAVS1 donor plasmid.
Cat # | Product Name | Description | Manual |
---|---|---|---|
pCas-Guide-AAVS1 | AAVS1 gRNA vector, validated AAVS1 targeting sequence cloned in pCas-Guide vector |
AAVS1 Donor Vectors
Transgene Knockin Vector Kits
AAVS1 Donor Controls
Cat # | Product Name | Promoter | Transgene/Selection Marker |
---|---|---|---|
pAAVS1-Cas9-Puro-DNR | CMV | Cas9/Puromycin | |
pAAVS1-Cas9-BSD-DNR | CMV | Cas9/Blasticidin | |
pAAVS1-RFP-DNR | CMV | tRFP/Puromycin | |
pAAVS1-mGFP-Puro-DNR | CMV | mGFP/Puromycin | |
pAAVS1-mRFP-Puro-DNR | CMV | mRFP/Puromycin | |
pAAVS1-mGFP-BSD-DNR | CMV | mGFP/Blasticidin | |
pAAVS1-mRFP-BSD-DNR | CMV | mRFP/Blasticidin | |
pAAVS1-EF1a-tGFP-puro-DNR | EF1a | tGFP/Puromycin | |
pAAVS1-EF1a-tGFP-BSD-DNR | EF1a | tGFP/Blasticidin | |
pAAVS1-EF1a-tRFP-puro-DNR | EF1a | tRFP/Puromycin | |
pAAVS1-EF1a-tRFP-BSD-DNR | EF1a | tRFP/Blasticidin | |
pAAVS1-EF1a-Luc-puro-DNR | EF1a | Luciferase/Puromycin | |
pAAVS1-EF1a-Luc-BSD-DNR | EF1a | Luciferase/Blasticidin |
More AAVS1 Products
ROSA26 Mouse Safe Harbor Knockin System via CRISPR
The ROSA26 site is the most popular genomic safe harbor site for mouse cell. It has been used to create more than a hundred mouse knock-in cell lines. The transgene inserted in this locus showed a constitutive and strong gene expression in mice.
OriGene has developed a series of ROSA26 CRISPR products to help researchers insert their gene of interest in the ROSA26 genomic locus through homologous recombination
This system needs two components:
- pCas-Guide-ROSA26, All-in-one CRISPR gRNA vector for ROSA26 targeting
- pAAVS1-Puro-DNR, donor vector for transgene integration, containing ROSA26 homologous arms (Gene of interest needs to be cloned)
Other CRISPR Products
Scheme of transgene knockin AAVS1/ROSA26 via CRISPR
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