Last data update: 24 January 2024 16:39 CET
Plasmid name: pDEST-N-HA (LMBP 7881)
| New search | Print data sheet |
| Price category: | Cat. 1 (cf. price list) |
| Status: | GeneCorner core plasmid |
| GeneCorner sequence: |
p7881.gb
(View with Genome Compiler) p7881.txt p7881.pdf |
| Sequence analysis results Genecorner: |
- |
| Cloned DNA: |
B gene of the control of cell death locus of the Escherichia coli F plasmid (ccdB, lethal gene) Influenza HA epitope encoding the haemagglutinin tagging peptide; N-terminal |
| Promoter: | Phaeodactylum tricornutum fucoxanthin chlorophyll a/c-binding protein B promoter (FcpB) Escherichia coli lac operon promoter Escherichia coli lac operon promoter; mutant (lacUV5) Phage T7 gene 10 promoter (T7g10) Phage T3 promoter |
| Ribosome binding site: |
Ribosome binding site (RBS) of the Escherichia coli lac Z gene (lacZ) |
| Terminator: | Phaeodactylum tricornutum fucoxanthin chlorophyll a/c-binding protein A terminator (FcpA) |
| Selection marker: | Ampicillin (amp) Chloramphenicol (cam) |
| Replicon: | Escherichia coli plasmid pMB1 origin Phage f1 origin |
| Host range: | Escherichia coli; use a ccdB-resistant strain for propagation Diatoms |
| Parental clone: | pBluescript KS+; pFCPBp-Sh ble; pKS-3HA |
| Further information: | The plasmid was constructed as follows: 1) The FcpB promoter and the FcpA terminator were PCR amplified from pFCPBp-Sh ble and ligated into the SacII-NotI sites and the EcoRI-KpnI sites of pBluescript KS+ respectively, generating pKS-FcpBpAt. 2) A DNA sequence encoding three repeats of the HA epitope was PCR amplified from pKS-3HA, and ligated into the NotI-BamHI sites of pKS-FcpBpAt, generating pKS-FcpBpAt-N-3HA. 3) The Gateway reading frame cassette A (rfA, from Invitrogen) was ligated into the SmaI site of pKS-FcpBpAt-N-3HA, generating pDEST-N-HA. pDEST-N-HA is a Gateway destination vector, containing the ccdB gene between the attenuator sites attR1 and attR2. It is meant for the creation of fusion proteins in diatoms, and contains three N-terminal HA epitopes. The nucleotide sequence of this plasmid corresponds with the EMBL Nucleotide Sequence Database accession number LT726868.1. The region downstream of the T7 promoter up to nucleotide position 1841 was sequenced by LMBP. The nucleotide sequence of the FcpB promoter corresponds with the EMBL Nucleotide Sequence Database accession number AF219942.1 except for an additional A nucleotide at position 1036, a C instead of T in position 1051 and a missing GC nucleotide pair between positions 1078 and 1079. The nucleotide sequence of the FcpA terminator corresponds with the EMBL Nucleotide Sequence Database accession number AF219942.1. |
| EMBL Accession number: | AF219942.1, view at EMBL, GenBank, DDBJ LT726868.1, view at EMBL, GenBank, DDBJ |
| Latest sequence update: | 18/12/2013 |
| Authenticity test: | Restriction enzyme pattern analysed at GeneCorner: EcoRI, HincII, KpnI and PvuII. |
| Class: | Recombinant plasmid |
| Type: | Plasmid |
| History of deposit: | This plasmid was deposited by Dr C. Bowler(1). (1) Institut de Biologie de l' Ecole Normale Supérieure (IBENS UMR8197 CNRS), Paris, France. |
| Plasmid reference: | Siaut et al., Gene 406 (2007), 23-35 [PMID: 17658702] |
| Restricted distribution: | - BCCM MTA |
| Distributed as: | H/P active culture or plasmid DNA |
| Host for distribution: | Escherichia coli K12xB DB3.1 |
| Host reference: | - |
| Related host reference: | Bernard et al., J. Mol. Biol. 226 (1992), 735-745 [PMID: 1324324] |
| Cultivation medium: | LB-Lennox + ampicillin (100 μg/ml) + chloramphenicol (34 μg/ml)* |
| Cultivation temperature: | 37°C |
| Biosafety level: | L1 |
| Cultivation remark: | *: selection of transformants on chloramphenicol; subsequent cultivation of a single colony in liquid medium with ampicillin. |
| Other culture collection numbers: | LMBP 07353 |
Refer in your Materials and Methods: |
pDEST-N-HA (LMBP 7881) is available at BCCM/GeneCorner. This plasmid was deposited by Dr C. Bowler and was published in Siaut et al., 2007. |
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.