Plasmid 12257: pWPXL

• Gene/insert name: None
• Fusion protein or tag: EGFP
• Vector backbone: pWPXL
  (Search Vector Database)
• Vector type: Mammalian Expression, Lentiviral, Cre/Lox
• Backbone size (bp): 10510
• Cloning site 5': MCS - see map
• Site destroyed during cloning: No
• Cloning site 3': MCS - see map
• Site destroyed during cloning: No
• 5' sequencing primer: See map
• Bacterial resistance(s) Ampicillin
• Growth strain(s) Stbl3
• Growth temperature (℃): 37
• Growth instructions: Use Stbl3 or HB101 to reduce chance of recombination. Grow at 37C
• High or low copy: High Copy
• Sequence: View sequences (2)
• Map: View map
• Supplemental document: Notes from Addgene (1)
• Principal Investigator: Didier Trono
• Terms and Licenses: MTA

Comments: 

pWPT or pWPXL can be used for constitutive transgene expression.
pWPXL contains the EF-1alpha promoter + intron that gives you high expression, as RNA loves to be spliced (it goes more efficiently out of the nucleus). pWPT contains only the EF-1alpha promoter

The loxP site in the 3'LTR is duplicated to the 5'LTR during reverse transcription in the target cells. This allows for removal (if necessary) of an integrated provirus by Cre.

Unique restriction sites at key positions will allow you to change promoter and transgene. PmeI is a popular site for cloning. You can also use PacI and SwaI.

Please note that ClaI in this vector is blocked by Dam methylation. This plasmid needs to be grown in a Dam- bacteria strain if you wish to use ClaI for cloning.

Packaging plasmids for Trono lab lentiviral vectors are also available at Addgene http://www.addgene.org/rnaitools

Please note that the full sequence for this plasmid is approximated and not fully verified. Please visit the Trono lab http://tronolab.epfl.ch for cloning strategies, protocols, publications, and more. See LentiWeb http://www.lentiweb.com for discussions on cloning strategies and protocols.