Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V013713 | pTRE3G-MCS-EGFP-3×FLAG-Tetone-Puro | In stock, instant shipping |
Buy one, get one free! |
Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.
Basic Vector Information
- Vector Name:
- pTRE3G-MCS-EGFP-3×FLAG-Tetone-Puro
- Antibiotic Resistance:
- Ampicillin
- Length:
- 7306 bp
- Type:
- Protein expression, Tetracycline inducible
- Replication origin:
- ori
- Host:
- Mammalian cells
- Selection Marker:
- Puro
- Promoter:
- TRE3G
- Growth Strain(s):
- DH10B
- Growth Temperature:
- 37℃
pTRE3G-MCS-EGFP-3×FLAG-Tetone-Puro vector Map
Plasmid Protocol
1. Centrifuge at 5,000×g for 5 min.
2. Carefully open the tube and add 20 μl of sterile water to dissolve the DNA.
3. Close the tube and incubate for 10 minutes at room temperature.
4. Briefly vortex the tube and then do a quick spin to concentrate the liquid at the bottom. Speed is less than 5000×g.
5. Store the plasmid at -20 ℃.
6. The concentration of plasmid re-measurement sometimes differs from the nominal value, which may be due to the position of the lyophilized plasmid in the tube, the efficiency of the re-dissolution, the measurement bias, and adsorption on the wall of the tube, therefore, it is recommended to transform and extract the plasmid before using it
General Plasmid Transform Protocol
1. Take one 100μl of the competent cells and thaw it on ice for 10min, add 2μl of plasmid, then ice bath for 30min, then heat-shock it at 42℃ for 60s, do not stir, and then ice bath for 2min.
2. Add 900μl of LB liquid medium without antibiotics, and incubate at 37℃ for 45min (30℃ for 1-1.5 hours) with 180rpm shaking.
3. Centrifuge at 6000rpm for 5min, leave only 100μl of supernatant to resuspend the bacterial precipitate and spread it onto the target plasmid-resistant LB plate.
4. Invert the plate and incubate at 37℃ for 14h, or at 30℃ for 20h.
5. Pick a single colony into LB liquid medium, add the corresponding antibiotics, incubate at 220rpm for 14h, and extract the plasmid according to the experimental needs and the instructions of the plasmid extraction kit.
pTRE3G-MCS-EGFP-3×FLAG-Tetone-Puro vector Sequence
LOCUS 62056_21425 7306 bp DNA circular SYN 01-JAN-1980
DEFINITION synthetic circular DNA.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 7306)
AUTHORS .
TITLE Direct Submission
FEATURES Location/Qualifiers
source 1..7306
/mol_type="other DNA"
/organism="synthetic DNA construct"
protein_bind 208..241
/label=loxP
/note="Cre-mediated recombination occurs in the 8-bp core
sequence (ATGTATGC) (Shaw et al., 2021)."
promoter 251..629
/label=TRE3G promoter
/note="3rd-generation Tet-responsive promoter that can be
activated by binding of Tet-On(R) 3G"
CDS 775..1491
/codon_start=1
/label=EGFP
/note="enhanced GFP"
/translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
EFVTAAGITLGMDELYK"
CDS 1540..1563
/codon_start=1
/label=FLAG
/note="FLAG(R) epitope tag, followed by an enterokinase
cleavage site"
/translation="DYKDDDDK"
polyA_signal 1581..1805
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
promoter 1901..2112
/label=EF-1-alpha core promoter
/note="core promoter for human elongation factor
EF-1-alpha"
LTR 2125..2393
/label=5' LTR (truncated)
/note="truncated 5' long terminal repeat (LTR) from human
T-cell leukemia virus (HTLV) type 1"
CDS 2427..3170
/codon_start=1
/label=Tet-On(R) 3G
/note="modified rtTA protein that binds tightly to
promoters containing the tet operator in the presence of
doxycycline"
/translation="MSRLDKSKVINSALELLNGVGIEGLTTRKLAQKLGVEQPTLYWHV
KNKRALLDALPIEMLDRHHTHSCPLEGESWQDFLRNNAKSYRCALLSHRDGAKVHLGTR
PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEEQEHQVAKEERETPTT
DSMPPLLKQAIELFDRQGAEPAFLFGLELIICGLEKQLKCESGGPTDALDDFDLDMLPA
DALDDFDLDMLSADALDDFDLDMLPG"
CDS 3171..3224
/codon_start=1
/label=T2A
/note="2A peptide from Thosea asigna virus capsid protein"
/translation="EGRGSLLTCGDVEENPGP"
CDS 3231..3827
/codon_start=1
/label=PuroR
/note="puromycin N-acetyltransferase"
/translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
polyA_signal 3950..4071
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
promoter complement(4350..4368)
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
primer_bind complement(4378..4394)
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
rep_origin 4536..4991
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
promoter 5017..5121
/label=AmpR promoter
CDS 5122..5979
/codon_start=1
/label=AmpR
/note="beta-lactamase"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
rep_origin 6153..6741
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
protein_bind 7029..7050
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 7065..7095
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 7103..7119
/label=lac operator
/note="The lac repressor binds to the lac operator to
inhibit transcription in E. coli. This inhibition can be
relieved by adding lactose or
isopropyl-beta-D-thiogalactopyranoside (IPTG)."
primer_bind 7127..7143
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
promoter 7164..7182
/label=T3 promoter
/note="promoter for bacteriophage T3 RNA polymerase"