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Gene/insert name: CAG promoters, GFP, tTR-KRAB, Tet-on
Insert size (bp): Unknown
Vector backbone: None (Search Vector Database)
Type of vector: Mammalian expression,Lentiviral
Backbone size (bp): 12877
5' Sequencing primer: See map (List of Sequencing Primers)
Bacteria resistance: Ampicillin
High or low copy: High Copy
Grow in standard E. coli @ 37C: No
Bacterial strain for growth and growth condition: Use Stbl3 or HB101 to reduce chance of recombination. Grow at 37C
Plasmid Provided In: Stbl3
Principal Investigator: Patrick Aebischer
Comments: Transgenes can be expressed from any RNA Pol II promoter as part of bicistronic unit comprising the KRAB-based
repressor; tetO sequences are inserted into the vector LTR. Tet-on and Tet-off versions rely on repressors that bind in
the absence or the presence of doxycycline, respectively. Addition of Pol III promoter-small hairpin RNA cassette allows
for drug-controllable RNA interference (Tet-on shRNA).
Cloning shRNA into pLVET, pLVCT, and pLVPT vectors: first clone shRNA into pLVTHM downstream of the tetO-H1 region.
Then cut pLVTHM containing your shRNA with MscI-FspI and clone the insert containing the 3'LTR to target plasmid opened
with MscI-FspI. FspI cuts into AmpR. This means for inverted clones AmpR will not be restored; after selection you will
be left with clones with the shRNA cassette and functional AmpR.
Article: A versatile tool for conditional gene expression and knockdown. Szulc J et al. (Nat Methods. 2006 Feb . 3(2):109-16. Pubmed)