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 insertational 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 donor vector ( Precision Shuttling System)
ROSA26 or AAVS1 diagram

gRNA/CRISPR cuts the genome, then the gene of interest (GOI) cassette will be knocked in via homologous recombination.

AAVS1 – Human Targeted Gene Insertion via CRISPR

ROSA26 – Mouse Targeted Gene Insertion via CRISPR


AAVS1 Safe Harbor knockin system via CRISPR

This system needs two components:

  1. pCas-Guide-AAVS1, All-in-one CRISPR vector for AAVS1 targeting
  2. pAAVS1-Puro-DNR, donor vector for transgene integration, containing the AAVS1 homologous arms (Gene of interest needs to be cloned)
AAVS1 Knockin Validation Data

AAVS1 gRNA Vector

Cat # Product Name Description Manual
GE100023 pCas-Guide-AAVS1 AAVS1 gRNA vector, validated AAVS1 targeting sequence cloned in pCas-Guide vector

AAVS1 Donor Vectors

Cat # Product Name Description Manual
GE100024 pAAVS1-Puro-DNR Donor vector with AAVS1 homologous arms cloned, ready for transgene cloning
GE100035 pAAVS1-BSD-DNR Donor vector with AAVS1 homologous arms cloned (blasticidin selection), ready for exogenous gene cloning (10 ug)
GE100046 pAAVS1-EF1a-Puro-DNR AAVS1 donor vector with AAVS1 homologous arms cloned (Puro selection), EF1a promoter, ready for exogenous gene cloning (10 ug)
GE100048 pAAVS1-EF1a-BSD-DNR AAVS1 donor vector with AAVS1 homologous arms cloned (blasticidin selection), EF1a promoter, ready for exogenous gene cloning (10 ug)

Transgene Knockin Vector Kits

Cat # Product Name Description Manual
GE100027 AAVS1 Transgene knockin vector kit Vector kit for targeted transgene insertion into AAVS1 locus, including GE100023, GE100024 and GE100003
GE100036 AAVS1 Transgene knockin kit (BSD) Vector kit for targeted transgene insertion into AAVS1 locus (blasticidin selection), including GE100023, GE100035 and GE100003
GE100047 AAVS1 Transgene Knockin kit (EF1a, Puro) Vector kit for targeted transgene insertion into AAVS1 locus (EF1a, Puro), including GE100023, GE100046 and GE100003
GE100049 AAVS1 Transgene Knockin kit (EF1a, BSD) Vector kit for targeted transgene insertion into AAVS1 locus (EF1a, blasticidin), including GE100023, GE100048 and GE100003

More AAVS1 Products

Cas9 insertion at AAVS1 locus
RFP AAVS1 donors
(AAVS1 positive control)
AAVS1-int5 PCR Primer Pairs

ROSA26 Mouse Safe Harbor Knockin System via CRISPR

This system needs two components:

  1. pCas-Guide-ROSA26, All-in-one CRISPR gRNA vector for ROSA26 targeting
  2. pAAVS1-Puro-DNR, donor vector for transgene integration, containing ROSA26 homologous arms (Gene of interest needs to be cloned)
ROSA26 Knockin validation data

ROSA26 gRNA Vector

Cat # Product Name Description Manual
GE100050 pCas-Guide-ROSA26 ROSA26 safe harbor gRNA vector, validated ROSA26 gRNA targeting sequence cloned in pCas-Guide vector

ROSA26 Donor Vector

Cat # Product Name Description Manual
GE100051 pROSA26-Puro-DNR ROSA26 donor vector with ROSA26 homologous arms cloned (Puro selection), ready for exogenous gene cloning (10 ug)

ROSA26 CRISPR Knockin Kit

Cat # Product Name Description Manual
GE100052 ROSA26 CRISPR transgene knockin kit (Puro) Vector kit for CRISPR targeted transgene insertion into ROSA26 locus in mice (Puro), including GE100050, GE100051 and GE100003

Other CRISPR Products

CRISPR Vectors
CRISPR Knockout Kits
Cas9 Nuclease
Cas9 Ab

Scheme of transgene knockin AAVS1/ROSA26 via CRISPR


AAVS1 transgene knockin diagram
ROSA26 transgene knockin diagram
基因表达相关资源