pJ607-CMV-GFP-2A-Cas9
(Plasmid #154830)

• Purpose: GFP-2A-Cas9 (CHO codon-optimized)
• Depositing Lab: Lise Marie Grav
• Publication: Sergeeva et al ACS Synth Biol. 2020 Aug 24. doi: 10.1021/acssynbio.0c00322.

Backbone

• Vector backbone: pJ607
• Backbone manufacturer: DNA2.0
• Backbone size w/o insert (bp): 2541
• Modifications to backbone: Amplified for USER cloning
• Vector type: Mammalian Expression, CRISPR
• Selectable markers: Hygromycin

Growth in Bacteria

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

Gene/Insert 1

• Gene/Insert name: GFP-2A-Cas9
• Insert Size (bp): 4941
• Mutation: Codon-optimized (Cricetulus griseus)
• Promoter: CMV

Cloning Information for Gene/Insert 1

• Cloning method: Unknown
• 5′ sequencing primer: unknown

Gene/Insert 2

• Gene/Insert name: HygR
• Insert Size (bp): 1026
• Promoter: SV40 promoter

Cloning Information for Gene/Insert 2

• Cloning method: Unknown
• 5′ sequencing primer: unknown

Resource Information

• Supplemental Documents:
  • pl0120_cled.gb

Terms and Licenses

• Academic/Nonprofit Terms:
  • UBMTA
• Industry Terms:
  • Not Available to Industry

Depositor Comments

This plasmid contains a codon-optimized Cas9 fused to eGFP for genome editing of Chinese hamster ovary (CHO) cells.

The plasmid was initially described in:
Grav LM et al. 2015 "One-step generation of triple knockout CHO cell lines using CRISPR/Cas9 and fluorescent enrichment" DOI: 10.1002/biot.201500027

A protocol describing the use of the plasmid for CRISPR/Cas9-mediated knockout in CHO cells:
Grav LM et al. 2017 "Application of CRISPR/Cas9 Genome Editing to Improve Recombinant Protein Production in CHO Cells" DOI: 10.1007/978-1-4939-6972-2_7

A protocol describing the use of the plasmid for CRISPR/Cas9-mediated targeted integration in CHO cells:
Sergeeva D et al. 2019 "CRISPR/Cas9 as a Genome Editing Tool for Targeted Gene Integration in CHO Cells". DOI: 10.1007/978-1-4939-9170-9_13

The plasmid was assembled by USER cloning.