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LMBP plasmid details

Last data update: 28 November 2020 04:24 CET

Plasmid name: pMG14 (LMBP 8465)

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Price category: Cat. 1 (cf. price list)
Status: GeneCorner core plasmid
GeneCorner sequence: p8465.gb (View with Genome Compiler)
p8465.txt
p8465.pdf
Sequence
analysis results
Genecorner:

-

Cloned DNA: Bacillus subtilis amylase gene (amyE); fragments
Strep-tag II; N-terminal
Promoter: Bacillus subtilis central glycolytic genes regulator promoter (cggR)
Escherichia coli lac operon promoter
Ribosome
binding site:
Ribosome binding site (RBS) of the Escherichia coli lac Z gene (lacZ)
Terminator: -
Selection marker: Ampicillin (amp)
Kanamycin (kan; tobramycin (tob))
Replicon: Escherichia coli plasmid pMB1 origin
Host range: Escherichia coli
Bacillus sp.; integrative
Parental clone: pMG9
Further information: The plasmid was constructed by PCR amplifying the B. subtilis cggR promoter as an EcoRI/PstI fragment and the B. subtilis gapA 5' UTR as a PstI/Acc65I fragment, and cloning them jointly into the EcoRI/Acc65I-opened pMG9 vector.
This plasmid is a modular E. coli–Bacillus shuttle vector, applicable for gene overexpression in B. subtilis via chromosomal integration.
The 5' UTR of B. subtilis gapA contains a transcriptional regulator. Genes can be integrated in another site by replacing the two amyE inserts as an AatII/XbaI fragment and SalI/NotI fragment respectively.
Furthermore, the modular architecture of the pMG plasmid family enables an easy exchange of all features like antibiotic resistance genes (EcoRI/SalI), promoters (EcoRI/PstI), 5' UTR (PstI/HindIII) and tags (HindIII/Acc65I) leading to a significant increase of applicabilities.
The EcoRI/SalI fragment of MG14 contains the kanamycin/tobramycin resistance gene aadD.
When cloning a fragment downstream from the lac promoter it may be advisable to use lacIq strains in order to prevent fortuitous expression of a possibly noxious polypeptide.
The nucleotide sequence of this plasmid corresponds with the EMBL Nucleotide Sequence Database accession number LT726805.1.
The nucleotide sequence of the B. subtilis fragments correspond with the EMBL Nucleotide Sequence Database accession number AP012496.1.
EMBL Accession number: AP012496.1, view at EMBL, GenBank, DDBJ
LT726805.1, view at EMBL, GenBank, DDBJ
Latest sequence update: 06/06/2013
Authenticity test: Restriction enzyme pattern analysed at GeneCorner: EcoRI/SalI, HaeII, HincII and NotI.
Class: Recombinant plasmid
Type: Plasmid
History of deposit: This plasmid was deposited by Dr. M. Gimpel(1) and Prof. Dr S. Brantl(1).
(1) AG Bakteriengenetik, FSU Jena, Jena, Germany
Plasmid reference: Gimpel et al., J. Microbiol. Methods 91 (2012), 312-317 [PMID: 22982324] [DOI: 10.1016/j.mimet.2012.09.003]
Restricted distribution: - BCCM MTA
- The depositor will be informed of the customer's identity upon release of a sample outside the depositor's department or outside the departments in which BCCM/GeneCorner is embedded, namely UGent-DBMB and VIB-IRC.
Distributed as: H/P active culture or plasmid DNA
Host for distribution: Escherichia coli K12 DH5α
Host reference: Focus 8 (1986), 9
Related host reference: Grant et al., Proc. Natl. Acad. Sci. U.S.A. 87 (1990), 4645-4649 [PMID: 2162051]
Hanahan, in 'DNA Cloning: A Practical Approach Volume I', IRL Press, Oxford (1985), 109-135 [ISSN/ISBN: 0947946187]
Hanahan, J. Mol. Biol. 166 (1983), 557-580 [PMID: 6345791]
Rodriguez-Quinones et al., Focus 15 (1993), 110-112
Woodcock et al., Nucleic Acids Res. 17 (1989), 3469-3478 [PMID: 2657660]
Cultivation medium: LB-Lennox + ampicillin (100 μg/ml) + kanamycin (50 μg/ml)
Cultivation temperature: 37°C
Biosafety level: L1
Cultivation remark: -
Other culture collection numbers: -

Note: Up-to-date, validated data are enclosed with the biological material. Nevertheless, these data are a snapshot at a given moment; further updates are always possible.

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