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LMBP plasmid details

Last data update: 28 November 2020 04:24 CET

Plasmid name: pMa58SAm2 (LMBP 1996)

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Price category: Cat. 1 (cf. price list)
Status: GeneCorner core plasmid
GeneCorner sequence: p1996.gb (View with Genome Compiler)
p1996.pdf
Sequence
analysis results
Genecorner:

-

Cloned DNA: Streptomyces avidinii streptavidin gene (SA); mutated sequence
Promoter: -
Ribosome
binding site:
-
Terminator: Phage fd terminator
Selection marker: Ampicillin (amp)
Replicon: Escherichia coli plasmid pMB1 origin
Phage f1 origin
Host range: Escherichia coli
Parental clone: pMa58; pMc58SA
Further information: The plasmid was derived from pMc58SA by introducing a BamHI site at the position of the termination codon of the Streptomyces avidinii streptavidin gene (SA), using the site-specific mutagenesis protocol of Stanssens et al. (1989).
The sequence of the streptavidin gene was obtained from Genbank (Release 55; Locus: STMSTRAVG) and shows, as compared to Dr Lieberman's sequence map of the parental clone pSA307, a few differences in the 5' and 3' untranslated regions of the gene.
The phasmid provides resistance to ampicillin but is sensitive to chloramphenicol.
More than one mutagenesis round is possible by switching from ampicillin resistance to chloramphenicol resistance.
pMa58SAm2 contains the replication origin of the single-stranded DNA phage f1, so that it can be rapidly switched between the plasmid and the phage mode (ssDNA) of replication. The latter requires infection with a helper phage M13KO7.
After mutagenesis, use mutS strains for primary transformation (e.g. sup(-) strain WK6mutS, sup(+) strain BMH71-18mutS); for segregation of possible mutants: sup(-) strains (e.g. WK6).
The nucleotide sequence of this plasmid corresponds with the EMBL Nucleotide Sequence Database accession number LT727616.1.
Other name of the plasmid is pSAB2.
EMBL Accession number: LT727616.1, view at EMBL, GenBank, DDBJ
Latest sequence update: 12/07/1990
Sequence detail:
Difference between the wt streptavidin gene and the mutated gene:

- wild type gene: ..C.GTT.CAG.CAG.TAG TCGCGTCC
                                  *
- mutated gene, making use of the following mutator oligonucleotide:

  C.GTT.CAG.CAG.GAT.CCGCGTCC
                ^
              BamHI

*: Termination codon of the wt streptavidin gene.
^: Last codon of the mature sequence of the mutated steptavidin gene.
Punctuation indicates reading frame.
Authenticity test: Restriction enzyme pattern analysed at GeneCorner: BamHI, BglI/SalI, EcoRI, EcoRI/HindIII and PstI.

The plasmid seems to be larger as compared to the compiled nucleotide sequence, the extra nucleotides being located in the vector part of the plasmid.
Class: Recombinant plasmid
Type: Plasmid
History of deposit: This plasmid was deposited by J. Viaene(1) and Prof. Dr E. Remaut(1).
(1) Department of Biomedical Molecular Biology, Ghent University, Ghent, Belgium
Plasmid reference: -
Restricted distribution: - BCCM MTA
Distributed as: H/P active culture or plasmid DNA
Host for distribution: Escherichia coli K12 MC1061
Host reference: Casadaban et al., J. Mol. Biol. 138 (1980), 179-207 [PMID: 6997493]
Related host reference: Brigé et al., Biochem. J. 394 (2006), 335-344 [PMID: 16293111]
Cultivation medium: LB-Lennox + ampicillin (100 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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