Basic Vector Information
In the Tet-Off expression system, a tetracycline-controlled transactivator protein (tTA), which is composed of the Tet repressor DNA binding protein (TetR) from the Tc resistance operon of Escherichia coli transposon Tn10 fused to the strong transactivating domain(sequence: PTDALDDFDLDML) of VP16 from Herpes simplex virus, regulates expression of a target gene that is under transcriptional control of a tetracycline-responsive promoter element (TRE). The TRE is made up of Tet operator (tetO) sequence concatemers fused to a minimal promoter. In the absence of Tc or Dox, tTA binds to the TRE and activates transcription of the target gene. In the presence of Tc or Dox, tTA cannot bind to the TRE, and expression from the target gene remains inactive. The Tet-On system is based on a reverse tetracycline-controlled transactivator, rtTA. Like tTA, rtTA is a fusion protein comprised of the TetR repressor and the VP16 transactivation domain; however, a four amino acid change(E19G, A56P, D148E, H179R) in the tetR DNA binding moiety alters rtTA's binding characteristics such that it can only recognize the tetO sequences in the TRE of the target transgene in the presence of the Dox effector. Thus, in the Tet-On system, transcription of the TRE-regulated target gene is stimulated by rtTA only in the presence of Dox.
- Vector Name:
- pTet-Off-Advanced
- Antibiotic Resistance:
- Ampicillin
- Length:
- 7140 bp
- Type:
- Mammalian Expression Vectors
- Replication origin:
- ori
- Source/Author:
- Clontech
- Selection Marker:
- Neomycin/G418(Geneticin)
- Copy Number:
- High copy number
- Promoter:
- SV40
pTet-Off-Advanced vector Vector Map
pTet-Off-Advanced vector Sequence
LOCUS pTet-Off-Advance 7140 bp DNA circular SYN 01-JAN-1980
DEFINITION Vector for expressing the tTA-Advanced transactivator in the
Tet-Off(R) Advanced system.
ACCESSION .
VERSION .
KEYWORDS pTet-Off-Advanced
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 7140)
AUTHORS Clontech
TITLE Direct Submission
REFERENCE 2 (bases 1 to 7140)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..7140
/mol_type="other DNA"
/organism="synthetic DNA construct"
enhancer 89..468
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 469..672
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
CDS 775..1518
/label=tTA-Advanced
/note="improved tetracycline-controlled transactivator"
polyA_signal 1531..1665
/label=SV40 poly(A) signal
/note="SV40 poly(A) signal"
/note="SV40 polyadenylation signal"
rep_origin complement(2385..2973)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(3147..4004)
/label=AmpR
/note="beta-lactamase"
promoter complement(4005..4109)
/label=AmpR promoter
polyA_signal complement(4222..4356)
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
CDS complement(4781..4801)
/label=SV40 NLS
/note="nuclear localization signal of SV40 (simian virus
40) large T antigen"
intron complement(4931..4996)
/label=small t intron
/note="SV40 (simian virus 40) small t antigen intron"
CDS complement(5419..6210)
/label=NeoR/KanR
/note="aminoglycoside phosphotransferase"
promoter complement(6571..6900)
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
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