AICSDP-35:AAVS1-mEGFP
(Plasmid #91565)

• Purpose: Homology arms and linker-mEGFP sequence for internal insertion of CAGGS-mEGFP at the AAVS1 safe harbor location in human cells as a reporter for cytoplasmic mEGFP
• Depositing Lab: Allen Institute for Cell Science
• Publication: Allen Institute for Cell Science Plasmids (unpublished)

Backbone

• Vector backbone: unknown
• Vector type: Mammalian Expression, CRISPR ; Donor Template
• Selectable markers: Puromycin

Growth in Bacteria

• Bacterial Resistance(s): Ampicillin, 100 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): DH5alpha
• Copy number: High Copy

Gene/Insert

• Gene/Insert name: AAVS1 Homology Arms with CAGGS driven mEGFP
• Alt name: CAGGS-mEGFP
• Species: H. sapiens (human)
• Insert Size (bp): 5944
• Entrez Gene: AAVS1 (a.k.a. AAV)
• Promoter: CAG

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ cloning site: unknown (unknown if destroyed)
• 3′ cloning site: unknown (unknown if destroyed)
• 5′ sequencing primer: unknown

Resource Information

• Articles Citing this Plasmid:
  • 3 References

Terms and Licenses

• Academic/Nonprofit Terms:
  • UBMTA
• Industry Terms:
  • Industry MTA

Depositor Comments

This plasmid has been used with locus-specific CRISPR/Cas9 to insert a CAGGS-mEGFP at the AAVS1 safe harbor locus in WTC human induced pluripotent stem cells by the Allen Institute for Cell Science. After insertion of the reporter using this donor template plasmid and CRISPR/Cas9 reagents, transfected cells may exhibit varying intensity levels of fluorescence, likely due to editing precision. To obtain cells of uniform intensity levels, see our protocol for fluorescence-assisted cell sorting and subcloning of transfected cells (https://www.allencell.org/instructional-videos-and-tutorials-for-cell-methods.html). Further, we recommend PCR-based assays for identifying precisely edited clones as previously described (https://www.molbiolcell.org/doi/abs/10.1091/mbc.e17-03-0209). For more information on the entire plasmid collection, please see https://www.addgene.org/allen-institute-cell-science/ .