3×Cherry Dam AID Smash
(Plasmid #227374)

• Purpose: For inducible expression and cytosolic localization of Dam
• Depositing Lab: David Pellman
• Publication: Papathanasiou et al Nature. 2023 Jul;619(7968):184-192. doi: 10.1038/s41586-023-06157-7. Epub 2023 Jun 7.

Backbone

• Vector backbone: pCW57.1
• Vector type: Mammalian Expression, Lentiviral
• Selectable markers: Puromycin

Growth in Bacteria

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

Gene/Insert

• Gene/Insert name: Dam DNA-adenine methyltransferase
• Species: H. sapiens (human)
• Tag / Fusion Protein:
  • mCherry (C terminal on insert)

Resource Information

• Supplemental Documents:
  • 3xCherryDamAIDSmash.gbk
• A portion of this plasmid was derived from a plasmid made by: David Root (Addgene plasmid #41393) and the van Steensel lab.

Terms and Licenses

• Academic/Nonprofit Terms:
  • UBMTA
  • Takara Bio Limited Use Label License (formerly Clontech)
• Industry Terms:
  • Not Available to Industry

Depositor Comments

The megaDam vector was generated by synthesizing (Genewiz) a sequence containing three copies of mCherry (based on the sequence from pHAGE-EFS-N22p-3XRFPnls; Addgene plasmid #75387) and the sequences encoding the mAID and SMASh degrons (from Lemmens, B. et al. DNA replication determines timing of mitosis by restricting CDK1 and PLK1 activation. Mol. Cell 71, 117–128.e3 (2018)). The Dam coding sequence was taken from TS52_pT_damonly (van Steensel lab). The sequence encoding the Dam–mCherry double-degron fusion was cloned and introduced into the lentiviral vector pCW57.1 (Addgene plasmid #41393, Addgene; gift from David Root).