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pUG27 vector (V002336) Gene synthesis in pUG27 backbone

Price Information

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V002336 pUG27 In stock, instant shipping

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

Basic Vector Information

Vector Name:
pUG27
Antibiotic Resistance:
Ampicillin
Length:
3850 bp
Type:
PCR template vector
Replication origin:
ori
Host:
Yeast
Source/Author:
Gueldener U, Heinisch J, Koehler GJ, Voss D, Hegemann JH.
Promoter:
TEF
Growth Strain(s):
Stbl3

pUG27 vector Map

Copy Sequence View Map
pUG273850 bp60012001800240030003600loxPHIS3MX6loxPT7 promoteroriAmpRAmpR promoterSP6 promoter

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.

pUG27 vector Sequence

LOCUS       40924_45478        3850 bp DNA     circular SYN 18-DEC-2018
DEFINITION  PCR template vector pUG27, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3850)
  AUTHORS   Gueldener U, Heinisch J, Koehler GJ, Voss D, Hegemann JH.
  TITLE     A second set of loxP marker cassettes for Cre-mediated multiple gene
            knockouts in budding yeast
  JOURNAL   Nucleic Acids Res. 30 (6), E23 (2002)
  PUBMED    11884642
REFERENCE   2  (bases 1 to 3850)
  AUTHORS   Gueldener U, Heinisch JH, Voss D, Koehler G, Hegemann JH.
  TITLE     Direct Submission
  JOURNAL   Submitted (23-AUG-2000) Institut fuer Mikrobiologie, 
            Heinrich-Heine-Universitaet, Universtitaetsstr. 1, Duesseldorf 
            40225, Germany
REFERENCE   3  (bases 1 to 3850)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 3850)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nucleic 
            Acids Res."; date: "2002"; volume: "30"; issue: "6"; pages: "E23"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (23-AUG-2000) Institut fuer Mikrobiologie, 
            Heinrich-Heine-Universitaet, Universtitaetsstr. 1, Duesseldorf 
            40225, Germany"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..3850
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    53..86
                     /label=loxP
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     gene            141..1341
                     /label=HIS3MX6
                     /note="yeast selectable marker encoding the S. pombe his5
                     gene, which corresponds to S. cerevisiae HIS3"
     misc_feature    1401..1434
                     /label=loxP site
                     /note="loxP site"
     protein_bind    complement(1401..1434)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     promoter        complement(1488..1506)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     rep_origin      complement(1764..2352)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(2526..3383)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(3384..3488)
                     /label=AmpR promoter
     promoter        3834..3850
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"