p416TEF vector (V009725) Gene synthesis in p416TEF backbone

COA reports Sequencing Results MSDS Download GenBank File(.gb)

Price Information

Cat No.	Plasmid Name	Availability	Buy one, get one free! (?)
V009725	p416TEF	In stock, instant shipping

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

Basic Vector Information

p416TEF is a low-copy yeast expression vector containing the TEF strong promoter to drive constitutive expression of exogenous genes. It optimises protein transport from the endoplasmic reticulum to the Golgi apparatus through overexpression of SEC16, thereby enhancing α-amylase secretion efficiency.

Vector Name:: p416TEF

Antibiotic Resistance:: Ampicillin

Length:: 5524 bp

Type:: Yeast centromeric expression vector

Replication origin:: ori

Host:: Yeast

Source/Author:: Mumberg D, Muller R, Funk M.

Promoter:: TEF1

Growth Strain(s):: DH10B

p416TEF vector Map

Copy Sequence View 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.

References

Goncharoff DK, Cabral R, Applebey SV, Pagadala M, Du Z, Li L. Defining Key Residues of the Swi1 Prion Domain in Prion Formation and Maintenance. Mol Cell Biol. 2021 Jun 23;41(7):e0004421. doi: 10.1128/MCB.00044-21. Epub 2021 Jun 23. PMID: 33941618; PMCID: PMC8224238.

p416TEF vector Sequence

LOCUS       40924_2787        5524 bp DNA     circular SYN 17-DEC-2018
DEFINITION  Yeast centromeric expression vector p416TEF, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5524)
  AUTHORS   Mumberg D, Muller R, Funk M.
  TITLE     Yeast vectors for the controlled expression of heterologous proteins
            in different genetic backgrounds
  JOURNAL   Gene 156 (1), 119-122 (1995)
  PUBMED    7737504
REFERENCE   2  (bases 1 to 5524)
  AUTHORS   Ralser M.
  TITLE     Direct Submission
  JOURNAL   Submitted (08-JAN-2007) Vertebrate Genomics, Max-Planck Institute 
            for Molecular Genetics, Ihnestrasse 73, Berlin 14195, Germany
REFERENCE   3  (bases 1 to 5524)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 5524)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Gene"; 
            date: "1995"; volume: "156"; issue: "1"; pages: "119-122"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (08-JAN-2007) Vertebrate Genomics, Max-Planck Institute for 
            Molecular Genetics, Ihnestrasse 73, Berlin 14195, Germany"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..5524
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    complement(70..573)
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an
                     autonomously replicating sequence"
     promoter        610..714
                     /label=AmpR promoter
     CDS             715..1572
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      1746..2334
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     protein_bind    2622..2643
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     promoter        2658..2688
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2696..2712
                     /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     2720..2736
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        2757..2775
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        2792..3191
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     primer_bind     3195..3211
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3245..3261)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     terminator      3264..3511
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     promoter        complement(3530..3548)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3558..3574)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     rep_origin      3719..4174
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     CDS             complement(4308..5108)
                     /codon_start=1
                     /label=URA3
                     /note="orotidine-5'-phosphate decarboxylase, required for
                     uracil biosynthesis"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     promoter        complement(5109..5329)
                     /label=URA3 promoter