mCherry-2xCL-YFP-Vpr
(Plasmid #105215)

• Purpose: Bi-functional Fluorescent reporter protein to visualize HIV-1 fusion and core-trafficking
• Depositing Lab: Gregory Melikyan
• Publication: Sood et al J Biol Chem. 2017 Oct 18. pii: M117.818088. doi: 10.1074/jbc.M117.818088.

Backbone

• Vector backbone: pcDNA3.1/Zeo(+)
• Backbone size w/o insert (bp): 5015
• Total vector size (bp): 6723
• Vector type: Mammalian 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: Vpr
• Alt name: Viral protein R
• Species: HIV-1
• Insert Size (bp): 291
• Promoter: CMV
• Tags / Fusion Proteins:
  • mCherry (N terminal on backbone)
  • eYFP (N terminal on insert)
  • PC2x (SQNY/IRKVL)

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ cloning site: HindIII - NotI - BamHI - XhoI (not destroyed)
• 3′ cloning site: NotI - BamHI - XhoI (not destroyed)
• 5′ sequencing primer: CMV-fwd
• 3′ sequencing primer: Vpr-Rev

Resource Information

• Article Citing this Plasmid:
  • 1 Reference

Terms and Licenses

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

Depositor Comments

The pMM310-BlamVpr plasmid reagent was obtained through the NIH AIDS Reagent Program, Division of AIDS, NIAID, NIH: pMM310(Cat#11444) from Dr. Michael Miller. The DNA sequences encoding for Beta-lactamase (Blam) was replaced with HindIII-mCherry-PC-NotI-PC-YFP-BamHI encoding sequences.

mCherry-pc2x-eYFPVpr plasmid reference:
Sood C, Francis AC, Desai TM, Melikyan GB.An improved labeling strategy enables automated detection of single-virus fusion and assessment of HIV-1 protease activity in single virions.J Biol Chem. 2017 Oct 18. pii: jbc.M117.818088. doi: 10.1074/jbc.M117.818088.

pMM310 plasmid reference:
Tobiume M, Lineberger JE, Lundquist CA, Miller MD, and Aiken C. Nef Does Not Affect the Efficiency of Human Immunodeficiency Virus Type 1 Fusion with Target Cells. J. Virology 77(19); 10645-50, 2003. [Abstract].

An in-depth description of a similar system: Cavrois M, De Noronha C, Greene WC. A sensitive and specific enzyme-based assay detecting HIV-1 virion fusion in primary T lymphocytes. Nat Biotechnol. 2002 Nov;20(11):1151-4. Epub 2002 Sep 30. [Abstract]