Last data update: 24 January 2024 16:39 CET
Plasmid name: pJBpn25lacZ (LMBP 3652)
| New search | Print data sheet |
| Price category: | Cat. 1 (cf. price list) |
| Status: | GeneCorner core plasmid |
| GeneCorner sequence: |
p3652.gb
(View with Genome Compiler) p3652.pdf |
| Sequence analysis results Genecorner: |
- |
| Cloned DNA: |
Escherichia coli lac Z gene (lacZ) Escherichia coli lac repressor gene (lacI) Broad-host-range plasmid RK2 trans-acting replication initiator protein DNA (trfA) |
| Promoter: | Phage T5 N25 promoter with two lac operator sequences (N25/O2) Transposon Tn5 neomycin resistance gene promoter (neo) Escherichia coli lac repressor promoter; mutant (lacI(q)) Phage T3 promoter |
| Ribosome binding site: |
Ribosome binding site (RBS) of the phage T7 gene 10 (T7g10) |
| Terminator: | Phage fd terminator Phage T7 gene 10 terminator (T7g10) Phage λ early leftward terminator (λ L) |
| Selection marker: | Tetracycline (tet) |
| Replicon: | Broad-host-range plasmid RK2 vegetative replication origin Broad-host-range plasmid RK2/RP4 origin of transfer (oriT) |
| Host range: | Escherichia coli Gram-negative bacterial strains |
| Parental clone: | pJBpn25LUC2; pLT10lacZT3 |
| Further information: | This IncP, broad-host-range plasmid was constructed as follows: the XbaI/SfiI fragment of pLT10lacZT3, containing the lacZ gene was inserted between the XbaI/SstI sites of pJBpn25LUC2, hereby replacing the luciferase gene. SfiI and SstI sites were blunted with T4 polymerase. pJBpn25lacZ contains the constitutive lacI(q) promoter in the opposite orientation relative to the lacZ cassette, avoiding transcription of the lacZ gene. The expression of the lacZ gene is controlled by the IPTG inducible T5 N25/O2 promoter. The promoter/operator element T5 N25/O2 consists of the highly efficient E. coli phage T5 N25 promoter and two lac operator sequences (Bujard et al. (1987)). Due to the presence of the lacI(q) (repressor)-N25/O2 (promoter/operator) cassette on the plasmid, regulated (IPTG inducible) expression should be possible in a wide range of Gram-negative hosts. The plasmid RK2 trfA gene encodes two polypeptide products P382 and P285 of 382 and 285 amino acids respectively, using a different initiation codon within the same reading frame. The resulting protein is essential for vegetative replication. Introduce the plasmid by electroporation, or through conjugation, using plasmid RK2/RP4 transfer functions (provided in trans by RK2/RP4, RK2/RP4-derived vectors, other replicons carrying RK2/RP4 transfer functions (e.g. pRK2073) or strains expressing the tra genes of RK2/RP4). The nucleotide sequence of this plasmid corresponds with the EMBL Nucleotide Sequence Database accession number LT727366.1. |
| EMBL Accession number: | LT727366.1, view at EMBL, GenBank, DDBJ |
| Latest sequence update: | 06/03/2001 |
| Authenticity test: | Restriction enzyme pattern analysed at GeneCorner: AvaIII(NsiI), AvaIII(NsiI)/EcoRI, BamHI, BamHI/EcoRI and SalI. Tests done on storage number 3894. |
| Class: | Recombinant plasmid |
| Type: | Plasmid |
| History of deposit: | This plasmid was deposited by Prof. Dr E. Remaut(1)(2). (1) Department for Molecular Biomedical Research, VIB, Ghent, Belgium (2) 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 + tetracycline (10 μg/ml) |
| Cultivation temperature: | 37°C |
| Biosafety level: | L1 |
| Other culture collection numbers: | - |
Refer in your Materials and Methods: |
pJBpn25lacZ (LMBP 3652) is available at BCCM/GeneCorner. This plasmid was deposited by Prof. Dr E. Remaut. |
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.