Last data update: 24 January 2024 16:39 CET
Plasmid name: pT10sOmpArPDI (LMBP 3057)
| New search | Print data sheet |
| Price category: | Cat. 1 (cf. price list) |
| Status: | GeneCorner core plasmid |
| GeneCorner sequence: |
p3057.gb
(View with Genome Compiler) p3057.pdf |
| Sequence analysis results Genecorner: |
- |
| Cloned DNA: |
Escherichia coli outer membrane protein A cDNA (ompA, tolG, GeneID 945571); signal sequence Rat protein disulfide isomerase cDNA (rPDI); mature sequence |
| Promoter: | Phage T7 gene 10 promoter (T7g10) |
| Ribosome binding site: |
Ribosome binding site (RBS) of the phage T7 gene 10 (T7g10) |
| Terminator: | Phage T7 gene 10 terminator (T7g10) |
| Selection marker: | Ampicillin (amp) |
| Replicon: | Escherichia coli plasmid pMB1 origin |
| Host range: | Escherichia coli |
| Parental clone: | pT10HB2Mms; pSE380rPDI |
| Further information: | The plasmid was constructed as follows: 1) pSE380rPDI was cleaved with BanI, filled in with Klenow DNA polymerase and further digested with BglII; 2) Subsequently, the BanI (nucleotide position 5563) - BglII fragment encoding rat protein disulfide isomerase was ligated to the SacI (blunted with T4 DNA polymerase) - BamHI vector fragment (obtained by ligation of the SacI-PstI fragment to the PstI-BamHI fragment) of pT10HB2Mms. This plasmid was used to produce high levels of enzymatically active rat protein disulfide isomerase in E. coli. For T7 driven expression use a strain containing a controllable T7 RNA polymerase gene: preferably a pT7POL plasmid (Mertens et al., Biotechnology 13 (1995), 175-179) or e.g. BL21(DE3). Proceed as follows: first transform the expression strain with the auxiliary plasmid, make the transformants competent again, and then transform with the expression plasmid. The nucleotide sequence of this plasmid corresponds with the EMBL Nucleotide Sequence Database accession number LT727429.1. In pT10sOmpArPDI, the complete signal sequence of the E. coli outer membrane protein II was fused in phase to the nucleotide sequence encoding the last two amino acids of the rat PDI signal sequence, followed by the mature rat PDI gene. The nucleotide sequence at this fusion was confirmed by DNA sequence analysis. The nucleotide sequence of the rPDI cDNA corresponds with the EMBL Nucleotide Sequence Database (Release 36; Accession number X02918). |
| EMBL Accession number: | X02918, view at EMBL, GenBank, DDBJ LT727429.1, view at EMBL, GenBank, DDBJ |
| Latest sequence update: | 22/06/1994 |
| Sequence detail: | Nucleotide sequence at the fusion between the ompA signal sequence and the mature rat PDI gene.
| |
Gln Ala|Gly Ala|Asp Ala
5' CAG.GCC|GGC.GCC|GAC.GCT 3'
*|-------|@
*: End of the ompA signal sequence.
-: Last two amino acids of the rat PDI signal sequence.
@: Start of the mature rat PDI gene.
Punctuation indicates reading frame. |
| Authenticity test: | Restriction enzyme pattern analysed at GeneCorner: AvaII, BalI, BglI, HindIII and NdeI. |
| Class: | Recombinant plasmid |
| Type: | Plasmid |
| History of deposit: | This plasmid was deposited by Dr K. De Sutter(1) (2). (1) Department for Molecular Biomedical Research, VIB, Ghent, Belgium (2) Department of Biomedical Molecular Biology, Ghent University, Ghent, Belgium |
| Plasmid reference: | De Sutter et al., Gene 141 (1994), 163-170 [PMID: 8163184] |
| Related plasmid reference: | Gibson et al., Biochemistry 45 (2006), 6363-6371 [PMID: 16700547] [DOI: 10.1021/bi060288q] |
| 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 |
| Other culture collection numbers: | - |
Refer in your Materials and Methods: |
pT10sOmpArPDI (LMBP 3057) is available at BCCM/GeneCorner. This plasmid was deposited by Dr K. De Sutter and was published in De Sutter et al., 1994. |
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.