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pTEF:ATP vector (V016758)

Price Information

Cat No. Plasmid Name Availability Buy one, get one free! (?)
V016758 pTEF:ATP In stock, 1 week for quality controls

Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.

Basic Vector Information

Vector Name:
pTEF:ATP
Antibiotic Resistance:
Ampicillin
Length:
8554 bp
Type:
Protein expression
Replication origin:
ori
Host:
Yeast
Selection Marker:
Neo/G418
Promoter:
TEF1
Growth Strain(s):
DH5a
Growth Temperature:
37℃

pTEF:ATP vector Map

Copy Sequence View Map
pTEF:ATP8554 bp4008001200160020002400280032003600400044004800520056006000640068007200760080008400CAP binding sitelac operatorM13 revCYC1 terminatormseCFPTEF1 promoterkanMXT7 promoterM13 fwdAmpR promoterAmpRori

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.

pTEF:ATP vector Sequence

LOCUS       62056_21110        8554 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 8554)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..8554
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    97..118
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     protein_bind    173..189
                     /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     197..213
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     terminator      complement(1205..1394)
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     CDS             complement(2579..3262)
                     /codon_start=1
                     /label=mseCFP
                     /note="monomeric super enhanced variant of CFP (Matsuda et
                     al., 2008)"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHRFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIK
                     AHFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAA"
     promoter        complement(3280..3684)
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     gene            3761..5117
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin
                     resistance (Wach et al., 1994)"
     promoter        complement(5726..5744)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5751..5767)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        6639..6743
                     /label=AmpR promoter
     CDS             6744..7601
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      7775..8363
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"