pMTL82151 vector (V004418)

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Price Information

Cat No.	Plasmid Name	Availability	Add to cart
V004418	pMTL82151	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 pMTL82151 is a clostridium shuttle plasmid. There are a series of plasmids from the pMTL80000 system. They differ in their Gram+ origins of replication: pMTL82151 (pBP1 from C. botulinum); pMTL83151 (pCB102 from C. butyricum); pMTL84151 (pCD6 from C. difficile); and pMTL85141 (pIM13 from Bacillus subtilis). Vector could be mobilisable via RP4/RK2 mating system, e.g. using E.coli strains S17.

Vector Name:: pMTL82151

Antibiotic Resistance:: Chloramphenicol

Length:: 5260 bp

Type:: Shuttle vector

Replication origin:: ori

Source/Author:: Heap JT, Pennington OJ, Cartman ST, Minton NP.

Copy Number:: High copy number

Growth Strain(s):: stbl3

Growth Temperature:: 37℃

pMTL82151 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

Heap JT, Pennington OJ, Cartman ST, Minton NP. A modular system for Clostridium shuttle plasmids. J Microbiol Methods. 2009;78(1):79-85.

pMTL82151 vector Sequence

LOCUS       Exported                5260 bp DNA     circular SYN 02-SEP-2024
DEFINITION  Exported.
ACCESSION   V004418
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5260)
  AUTHORS   Heap JT, Pennington OJ, Cartman ST, Minton NP.
  TITLE     A modular system for Clostridium shuttle plasmids
  JOURNAL   J. Microbiol. Methods 78 (1), 79-85 (2009)
  PUBMED    19445976
REFERENCE   2  (bases 1 to 5260)
  AUTHORS   Pennington OJ, Heap JT, Cartman ST, Minton NP.
  TITLE     Direct Submission
  JOURNAL   Submitted (02-MAR-2009) Centre for Biomolecular Sciences, The 
            University of Nottingham, University Park, Nottingham NG7 2RD, 
            United Kingdom
REFERENCE   3  (bases 1 to 5260)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 5260)
  TITLE     Direct Submission
REFERENCE   5  (bases 1 to 5260)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "J.
            Microbiol. Methods"; date: "2009"; volume: "78"; issue: "1"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (02-MAR-2009) Centre for Biomolecular Sciences, The University of 
            Nottingham, University Park, Nottingham NG7 2RD, United Kingdom"
COMMENT     SGRef: number: 3; type: "Journal Article"
COMMENT     SGRef: number: 4; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..5260
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     source          2801..2806
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      9..62
                     /note="terminator derived from CD0164 gene of Clostridium 
                     difficile"
                     /regulatory_class="terminator"
     primer_bind     63..79
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(199..215)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     regulatory      363..404
                     /note="terminator derived from fdx gene of Clostridium 
                     pasteurianum"
                     /regulatory_class="terminator"
     CDS             925..1980
                     /codon_start=1
                     /gene="repA"
                     /product="plasmid replication protein"
                     /label=repA
                     /protein_id="ACR43886.1"
                     /translation="MTNYNQKNENKQEVKTALEKCTDKKRKNPRFTSYIAPLTTKKNIE
                     RTSTCGDYLFMLSDADLEHFKLHKGNFCGNRFCPMCSWRLACKDSLEISILMEHLRKEE
                     NKEFIFLTLTTPNVKSYDLNYSIKQYNKSFKKLMERKEVKDITKGYIRKLEVTYQKEKY
                     ITKDLWKIKKDYYQKKGLEIGDLEPNFDTYNPHFHVVIAVNKSYFTDKNYYINRERWLE
                     LWKFATKDDSITQVDVRKAKINDYKEVYELAKYSAKDTDYLISRPVFEIFYKALKGKQV
                     LVFSGFFKDAHKLYKQGKLDVYKKKDEIKYVYIVYYNWCKKQYEKTRIRELTEDEKEEL
                     NQDLIDEIEID"
     gene            925..1980
                     /gene="repA"
                     /label=repA
     CDS             complement(2071..2574)
                     /codon_start=1
                     /product="putative plasmid replication protein"
                     /label=putative plasmid replication protein
                     /note="orf2"
                     /protein_id="ACR43887.1"
                     /translation="MKFKKIILYIFIVIIPLSITGCSINQITTPVPKNYLDNIVKKYVG
                     PEGVQPSFGGKTFYDYKIIDSEKSKNIIKIYLYYIGQEYYIEDDKLVEGTGSASFATVT
                     IKKENNNYKLVSYEVPKSPAQEDTVKIFPKSVRKKAIHANTPSLKNIEKEAETYFKKEL
                     KTKN"
     CDS             2904..3524
                     /gene="catP"
                     /label=Chloramphenicol acetyltransferase
                     /note="Chloramphenicol acetyltransferase from Clostridium 
                     perfringens. Accession#: P26826"
     rep_origin      3704..4292
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     oriT            4613..4722
                     /label=oriT
                     /note="incP origin of transfer"
     CDS             4755..5123
                     /label=traJ
                     /note="oriT-recognizing protein"

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