pNeDaKO-Hyg
(Plasmid #16414)

• Depositing Lab: Bert Vogelstein
• Publication: Kohli et al Nucleic Acids Res. 2004 . 32(1):e3.

Backbone

• Vector backbone: pBluescript SK+
• Backbone manufacturer: Stratagene
• Backbone size w/o insert (bp): 3000
• Vector type: Mammalian Expression, Cre/Lox
• Selectable markers: Hygromycin, Zeocin

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: PGK-HygR & ZeoR

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ cloning site: BstXI (unknown if destroyed)
• 3′ cloning site: KpnI (unknown if destroyed)
• 5′ sequencing primer: mPGK-F
• 3′ sequencing primer: T7

Terms and Licenses

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

Depositor Comments

Part of vector system to facilitate and simplify the use of rAAV for creating KOs in human cells. The other vectors for this system, pAAV-MCS, pAAV-RC, and pHelper, can be purchased from Stratagene.

The sequence information is the sequence between linkers A and B.

See author's map and article for more information. In the author's map, restriction sites for the generation of pNeDaKO fragments used for fusion PCR are shown. A and B refer to linkers A and B; yellow arrows: loxP sites; PGK: phosphoglycerate kinase eukaryotic promoter; Hyg: hygromycin resistance gene; EM 7: EM7 prokaryotic promoter; Zeo: zeomycin resistance gene; Amp: ampicillin resistance gene.

Note that though the Hyg-R gene has a point mutation, the Vogelstein lab has used this construct to for Hyg selection extensively.