pUltra-hot
(Plasmid #24130)

• Purpose: (Empty Backbone) 3rd generation Lentiviral vector for bi-cistronic expression of mCherry and the gene of interest
• Depositing Lab: Malcolm Moore
• Publication: pUltra series (unpublished)

Backbone

• Vector backbone: pRRL
• Backbone manufacturer: Didier Trono
• Backbone size (bp): 7000
• Vector type: Mammalian Expression, Lentiviral

Growth in Bacteria

• Bacterial Resistance(s): Ampicillin, 100 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): Stbl3
• Growth instructions: Stbl3 or SC110 (See below**)
• Copy number: Unknown

Gene/Insert

• Gene/Insert name: none
• Tag / Fusion Protein:
  • mCherry (N terminal on backbone)

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ cloning site: XbaI/NheI (not destroyed)
• 3′ cloning site: BamHI/BcII (not destroyed)
• 5′ sequencing primer: hUBCpro-F

Resource Information

• Addgene Notes:
  • Addgene Diagnostic Digest
• Articles Citing this Plasmid:
  • 31 References

Terms and Licenses

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

Depositor Comments

pUltra was cloned by Yildirim Dogan ()

This is a 3rd generation Lentiviral vector with an internal Ubiquitinc Promoter.

By cloning into the compatible cloning sites* (XbaI and BamHI) downstream of mCherry-P2A, you get a bi-cistronic expression of mCherry and the gene of interest. You can clone a second gene of interest into the NheI/bclI**donwstream of mCherry-P2A-gene1-T2A and get a multi-cistronic expression of all three genes.

* compatible cloning site (ccs): you can pcr amplifiy your gene of interest with a forward primer including one of these cutting sites: SpeI or NheI or XbaI and a reverse primer with one these cutting sites: BglII or BamHI or bclI. The PCR product is now compatible with the first site as well the 2nd site. "You can mix and match".

** for bclI the vector has to be amplified by dam-methylation defective E coli strains (e.g. SC110 Stratagene).

Further advantage of the vector is, that you can simultaneously do RNAi by cloning H1-shRNA cassettes into the unique SnaBI site in the 3´-LTR. Plus while integration the RNAi cassette gets doubled, since the 5´LTR is dublicated while reverse transcripton from the 3´-LTR

P2A: 3971-4033 and T2A is between bp 4049-4111