pLenti-Tet-MKK6(DD)-Puro
(Plasmid #86094)

• Purpose: Tet/Dox inducible (TetR) constitutively active mutant MKK6/MAP2K6 in lentiviral vector
• Depositing Lab: Cindy Miranti
• Publication: Frank et al J Cell Sci. 2017 Apr 26. pii: jcs.197152. doi: 10.1242/jcs.197152.

Backbone

• Vector backbone: pLenti CMV/TO Puro DEST (670-1)
• Backbone manufacturer: Eric Campeau
• Backbone size w/o insert (bp): 8000
• Total vector size (bp): 9194
• Modifications to backbone: L/R recombination to clone in MKK6-DD insert.
• 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: High Copy

Gene/Insert

• Gene/Insert name: MKK6
• Alt name: MAP2K6
• Alt name: MEK6
• Alt name: MKK6-DD, constitutively active mutant
• Species: H. sapiens (human)
• Insert Size (bp): 1005
• Mutation: Serine 207 and Threonine 211 both changed to Aspartic Acid (D)
• GenBank ID: NM_002758.3
• Entrez Gene: MAP2K6 (a.k.a. MAPKK6, MEK6, MKK6, PRKMK6, SAPKK-3, SAPKK3)
• Promoter: CMV/TetO
• Tag / Fusion Protein:
  • Myc tag (N terminal on insert)

Cloning Information

• Cloning method: Gateway Cloning
• 5′ sequencing primer: TGGGCGGTAGGCGTGTACG
• 3′ sequencing primer: GCAGCGTATCCACATAGCG

Resource Information

• Supplemental Documents:
  • pLenti-Tet-MKK6(DD)-Puro.gb
• A portion of this plasmid was derived from a plasmid made by: MKK6-DD gene construct originally made in the lab of Angel Nebreda. Citation in comments.
• Article Citing this Plasmid:
  • 1 Reference

Terms and Licenses

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

Depositor Comments

Note: The MKK6 insert in this plasmid has a S2A mutation compared to wild type, but the depositor has confirmed that it does not affect plasmid function.

For proper use, cell line must first be engineered to express the Tetracycline Repressor (TetR). For example: Addgene plasmid #17492. (Not compatible with rtTA system)

Source of MKK6-DD mutant construct:

Alonso, G., Ambrosino, C., Jones, M. and Nebreda, A.R., 2000. Differential activation of p38 mitogen-activated protein kinase isoforms depending on signal strength. Journal of Biological Chemistry, 275(51), pp.40641-40648.