14-3-3 KO vector
(Plasmid #16551)

• Depositing Lab: Bert Vogelstein
• Publication: Chan et al Nature. 1999 Oct 7. 401(6753):616-20.

Backbone

• Vector backbone: pBluescript
• Backbone manufacturer: Stratagene
• Backbone size w/o insert (bp): 3000
• Vector type: Bacterial Expression

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: 14-3-3 knockout
• Alt name: 14-3-3 sigma
• Species: H. sapiens (human)
• Entrez Gene: SFN (a.k.a. YWHAS)

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ cloning site: HindIII (not destroyed)
• 3′ cloning site: HindIII (not destroyed)
• 5′ sequencing primer: T7
• 3′ sequencing primer: T3

Terms and Licenses

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

Depositor Comments

A BAC clone was digested with BamHI, and two fragments, one 4.7 kilobases (kb) and the second 3.8 kb, were used to construct the 5' and 3' arms of the targeting vector, respectively. The 4.7-kb subclone contained the region immediately 5' of the initiating codon of the 14-3-3sigma coding region. The 3.8-kb subclone contained a region beginning 900 bp distal to the initiating codon. A synthetic EcoRV site plus the sequence 5'-CGTGGAGAGGGACT GGCAG-3', derived from a region of the 14-3-3sigma locus deleted by the targeting construct, were ligated to the 5' end of a geneticin-resistance gene. This product was placed into the pBluescript plasmid. To facilitate knockout of both alleles, loxP sites surrounding the geneticin-resistance gene were incorporated into the vector.