Last data update: 24 January 2024 16:39 CET
Plasmid name: pENkexhGNTIf (LMBP 3946)
| New search | Print data sheet |
| Price category: | Cat. 1 (cf. price list) |
| Status: | GeneCorner non-core plasmid |
| Depositor's sequence: | p3946.gb |
| Sequence analysis results Genecorner: |
- |
| Cloned DNA: |
Trichoderma reesei cellobiohydrolase I gene (cbhI); missing the cellulose binding domain Human N-acetylglucosaminyltransferase I cDNA (GlcNAc-TI, MGAT1); mature sequence with KEX2 protease cleavage site |
| Promoter: | Trichoderma reesei cellobiohydrolase I promoter (cbhI) |
| Ribosome binding site: |
- |
| Terminator: | Trichoderma reesei cellobiohydrolase I terminator (cbhI) |
| Selection marker: | Ampicillin (amp) |
| Replicon: | Escherichia coli plasmid pMB1 origin |
| Host range: | Escherichia coli Fungi |
| Parental clone: | pEN208; pUChGNTI1 |
| Further information: | The plasmid was constructed as follows: i) The StuI site of pEN208, located in the unknown region of the cbhI promoter, was eliminated by inserting a BglII linker, creating the pENBglII vector. ii) an oligo, containing a KEX2 protease cleaving site and a StuI site, was inserted between the EagI and BclI opened pENBglII vector, resulting in pEN208Kex2, iii) the Eco47III-HindII fragment of pUChGNTI1, containing the human N-acetylglucosaminyltransferase I cDNA (GlcNAc-TI) missing its signal sequence, was then inserted into the StuI-BclI (filled in) opened pEN208Kex2 vector. This plasmid was developed for secretion of truncated human GlcNAc-TI (Not experimentally verified). Selection of Trichoderma transformants is only possible upon cotransformation with a plasmid containing a selection marker. Since the nucleotide sequence of the 5' end of the cbh1 promoter ((+-) 600 bp) is not known, there is uncertainty as to the cutting frequency of the restriction enzymes indicated. The nucleotide sequence of the human GlcNAc-TI cDNA corresponds with the Genbank accession number M55621.1. |
| EMBL Accession number: | M55621.1, view at EMBL, GenBank, DDBJ |
| Latest sequence update: | 09/10/2001 |
| Sequence detail: | Oligo sequence:
|
5' TAC.GGC.CGC.CCC.ATG.TAC.AAG.AGG|CCT.GAT.ATC.GCG.GCC.GCT.TAA.TTGATCA 3'
EagI StuI EcoRV NotI --- BclI
|
|: KEX2 protease cleaving site. |
| Authenticity test: | The plasmid still needs to be subjected to the authenticity test. |
| Class: | Recombinant plasmid |
| Type: | Plasmid |
| History of deposit: | This plasmid was deposited by Prof. Dr N. Callewaert(1) (2). (1) Department for Molecular Biomedical Research, VIB, Ghent, Belgium (2) Department of Biomedical Molecular Biology, Ghent University, Ghent, Belgium |
| Plasmid reference: | - |
| Related plasmid reference: | Maras et al., FEBS Lett. 452 (1999), 365-370 [PMID: 10386623] |
| 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: | - |
Refer in your Materials and Methods: |
pENkexhGNTIf (LMBP 3946) is available at BCCM/GeneCorner. This plasmid was deposited by Prof. Dr N. Callewaert . |
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.