pDS132 vector (V007924)

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Price Information

Cat No.	Plasmid Name	Availability	Add to cart
V007924	pDS132	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

The pDS132 is a suicide plasmid. It can be easily used to introduce various types of mutations into different genetics backgrounds: removal of IS elements, introduction of point mutations or deletions.

Vector Name:: pDS132

Antibiotic Resistance:: Chloramphenicol

Length:: 5286 bp

Type:: Cloning vector

Replication origin:: R6K γ ori

Source/Author:: Philippe N, Alcaraz JP, Coursange E, Geiselmann J, Schneider D.

Promoter:: sacB

pDS132 vector Map

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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

Philippe N, Alcaraz JP, Coursange E, Geiselmann J, Schneider D. Improvement of pCVD442, a suicide plasmid for gene allele exchange in bacteria. Plasmid. 2004 May;51(3):246-55.

pDS132 vector Sequence

LOCUS       40924_15185        5286 bp DNA     circular SYN 17-DEC-2018
DEFINITION  Cloning vector pDS132, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5286)
  AUTHORS   Philippe N, Alcaraz JP, Coursange E, Geiselmann J, Schneider D.
  TITLE     Improvement of pCVD442, a suicide plasmid for gene allele exchange 
            in bacteria
  JOURNAL   Plasmid 51 (3), 246-255 (2004)
  PUBMED    15109831
REFERENCE   2  (bases 1 to 5286)
  AUTHORS   Philippe N, Alcaraz JP, Coursange E, Geiselmann J, Schneider D.
  TITLE     Direct Submission
  JOURNAL   Submitted (27-NOV-2003) LAPM, UMR 5163, BP53 CERMO, Grenoble 38041, 
            France
REFERENCE   3  (bases 1 to 5286)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 5286)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Plasmid"; 
            date: "2004"; volume: "51"; issue: "3"; pages: "246-255"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (27-NOV-2003) LAPM, UMR 5163, BP53 CERMO, Grenoble 38041, France"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..5286
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(4..392)
                     /direction=LEFT
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K;
                     requires the R6K initiator protein pi for replication"
     CDS             complement(1013..1381)
                     /label=traJ
                     /note="oriT-recognizing protein"
     oriT            complement(1414..1523)
                     /direction=LEFT
                     /label=oriT
                     /note="incP origin of transfer"
     CDS             complement(2492..3148)
                     /label=CmR
                     /note="chloramphenicol acetyltransferase"
     CDS             complement(3249..4667)
                     /label=SacB
                     /note="secreted levansucrase that renders bacterial growth 
                     sensitive to sucrose"
     promoter        complement(4668..5113)
                     /label=sacB promoter
                     /note="sacB promoter and control region"
     misc_feature    5135..5208
                     /note="multiple cloning site 
                     (XbaI/SalI/PstI/SalI/PstI/SalI/XbaI/SphI)"
     protein_bind    complement(5239..5255)
                     /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)."