pDM4 vector (V007969)

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Price Information

Cat No.	Plasmid Name	Availability	Add to cart
V007969	pDM4	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

pDM4 plasmid is a suicidal vector, which needs the strain containing pir gene. It is a derivative of plasmid pNQ705, a chloramphenicol-resistant derivative of the pGP704 vector that contains the oriV replicon originally from the plasmid R6K.

Vector Name:: pDM4

Antibiotic Resistance:: Chloramphenicol

Length:: 7104 bp

Type:: Cloning vector

Replication origin:: R6K γ ori

Source/Author:: Mou X, Spinard EJ, Driscoll MV, Zhao W, Nelson DR.

Selection Marker:: SacB

Copy Number:: Low copy number

Promoter:: sacB

Growth Strain(s):: S17-1λpir

Growth Temperature:: 37℃

pDM4 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

Lau TV, Puah SM, Tan JMA, Merino S, Puthucheary SD, Chua KH. Flagellar motility mediates biofilm formation in Aeromonas dhakensis. Microb Pathog. 2023 Apr;177:106059.

pDM4 vector Sequence

LOCUS       40924_14860        7104 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pDM4, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7104)
  AUTHORS   Mou X, Spinard EJ, Driscoll MV, Zhao W, Nelson DR.
  TITLE     H-NS Is a Negative Regulator of the Two Hemolysin/Cytotoxin Gene 
            Clusters in Vibrio anguillarum
  JOURNAL   Infect. Immun. 81 (10), 3566-3576 (2013)
  PUBMED    23836825
REFERENCE   2  (bases 1 to 7104)
  AUTHORS   Mou X, Spinard EJ, Driscoll MV, Zhao W, Nelson DR.
  TITLE     Direct Submission
  JOURNAL   Submitted (19-MAR-2013) Cell and Molecular Biology, University of 
            Rhode Island, 120 Flagg Rd, Kingston, RI 02881, USA
REFERENCE   3  (bases 1 to 7104)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 7104)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Infect. 
            Immun."; date: "2013"; volume: "81"; issue: "10"; pages: "3566-3576"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (19-MAR-2013) Cell and Molecular Biology, University of Rhode 
            Island, 120 Flagg Rd, Kingston, RI 02881, USA"
COMMENT     SGRef: number: 3; type: "Journal Article"
COMMENT     ##Assembly-Data-START##
            Sequencing Technology :: Sanger dideoxy sequencing 
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..7104
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     oriT            623..732
                     /label=oriT
                     /note="incP origin of transfer"
     CDS             765..1133
                     /codon_start=1
                     /label=traJ
                     /note="oriT-recognizing protein"
                     /translation="MADETKPTRKGSPPIKVYCLPDERRAIEEKAAAAGMSLSAYLLAV
                     GQGYKITGVVDYEHVRELARINGDLGRLGGLLKLWLTDDPRTARFGDATILALLAKIEE
                     KQDELGKVMMGVVRPRAEP"
     CDS             1192..1818
                     /codon_start=1
                     /product="conjugal transfer relaxase TraI"
                     /label=conjugal transfer relaxase TraI
                     /protein_id="AGO64135.1"
                     /translation="MRSIKKSDFAELVKYITDEQGKTERLGHVRVTNCEANTLPAVMAE
                     VMATQHGNTRSEADKTYHLLVSFRAGEKPDAETLRAIEDRICAGLGFAEHQRVSAVHHD
                     TDNLHIHIAINKIHPTRNTIHEPYRAYRALADLCATLERDYGLERDNHETRQRVSENRA
                     NDMERHAGVESLVGWIRPRCVRRRGSEDQQFNLLIVRTKLSCFTY"
     rep_origin      1754..2142
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K;
                     requires the R6K initiator protein pi for replication"
     protein_bind    2177..2193
                     /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)."
     promoter        2574..2676
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding
                     chloramphenicol acetyltransferase"
     CDS             2677..3333
                     /codon_start=1
                     /label=CmR
                     /note="chloramphenicol acetyltransferase"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     promoter        3753..4198
                     /label=sacB promoter
                     /note="sacB promoter and control region"
     CDS             4199..5617
                     /codon_start=1
                     /label=SacB
                     /note="secreted levansucrase that renders bacterial growth 
                     sensitive to sucrose"
                     /translation="MNIKKFAKQATVLTFTTALLAGGATQAFAKETNQKPYKETYGISH
                     ITRHDMLQIPEQQKNEKYQVPEFDSSTIKNISSAKGLDVWDSWPLQNADGTVANYHGYH
                     IVFALAGDPKNADDTSIYMFYQKVGETSIDSWKNAGRVFKDSDKFDANDSILKDQTQEW
                     SGSATFTSDGKIRLFYTDFSGKHYGKQTLTTAQVNVSASDSSLNINGVEDYKSIFDGDG
                     KTYQNVQQFIDEGNYSSGDNHTLRDPHYVEDKGHKYLVFEANTGTEDGYQGEESLFNKA
                     YYGKSTSFFRQESQKLLQSDKKRTAELANGALGMIELNDDYTLKKVMKPLIASNTVTDE
                     IERANVFKMNGKWYLFTDSRGSKMTIDGITSNDIYMLGYVSNSLTGPYKPLNKTGLVLK
                     MDLDPNDVTFTYSHFAVPQAKGNNVVITSYMTNRGFYADKQSTFAPSFLLNIKGKKTSV
                     VKDSILEQGQLTVNK"
     CDS             complement(5758..6135)
                     /codon_start=1
                     /product="insertion element IS1 1/5/6 protein insB"
                     /label=insertion element IS1 1/5/6 protein insB
                     /protein_id="AGO64138.1"
                     /translation="MDEQWGYVGAKSRQRWLFYAYDSLRKTVVAHVFGERTMATLGRLM
                     SLLSPFDVVIWMTDGWPLYESRLKGKLHVISKRYTQRIERHNLNLRQHLARLGRKSLSF
                     SKSVELHDKVIGHYLNIKHYQ"
     CDS             6413..6649
                     /codon_start=1
                     /product="beta-lactamase"
                     /EC_number="3.5.2.6"
                     /label=beta-lactamase
                     /protein_id="AGO64139.1"
                     /translation="MEADKVAGPLLRSALPAGWFIADKSGAGERGSRGIIAALGPDGKP
                     SRIVVIYTTGSQATMDERNRQIAEIGASLIKHW"