pGLS3-5xHRE-PHLDA3(wt)
(Plasmid #72557)

• Purpose: Hypoxia induced mutant PHLDA3. Contains the HRE from VEGF sequence (5 repeats) upstream of PHLDA3.
• Depositing Lab: Ester Hammond
• Publication: Leszczynska et al J Clin Invest. 2015 Jun;125(6):2385-98. doi: 10.1172/JCI80402. Epub 2015 May 11.

Backbone

• Vector backbone: pGL3-Basic
• Backbone manufacturer: Promega
• Modifications to backbone: PHLDA3 was cloned so that it replaced p53 from pGLS3-5xHRE-p53
• Vector type: Mammalian Expression
• Selectable markers: Neomycin (select with G418)

Growth in Bacteria

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

Gene/Insert

• Gene/Insert name: PHLDA3
• Alt name: pleckstrin homology-like domain, family A, member 3
• Species: H. sapiens (human)
• Entrez Gene: PHLDA3 (a.k.a. TIH1)
• Promoter: E1b minimal
• Tag / Fusion Protein:
  • 5XHRE (N terminal on insert)

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ cloning site: HindIII (not destroyed)
• 3′ cloning site: XbaI (not destroyed)
• 5′ sequencing primer: E1B F: 5’-agatctggtaggcgtgtacgg-3’
• 3′ sequencing primer: pGLS3 R: 5’-gtatcttatcatgtctgctcg-3’

Terms and Licenses

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

Depositor Comments

5xHRE-wtp53 plasmid was used as a backbone, where p53 sequence was cut out with HindIII and XbaI enzymes.
PHLDA3 was subsequently cloned in, so PHLDA3 cloning site contains:
KpnI – 5xHRE from VEGF – BglII – E1b minimal promoter – HindIII – kozak –PHLDA3 – XbaI