Last data update: 24 January 2024 16:39 CET
Plasmid name: pPP1MFmds1 (LMBP 4426)
| New search | Print data sheet |
| Price category: | Cat. 2 (cf. price list) |
| Status: | GeneCorner core plasmid |
| GeneCorner sequence: |
p4426.gb
(View with Genome Compiler) p4426.txt p4426.pdf |
| Sequence analysis results Genecorner: |
Sanger: ef32769308.fasta Sanger: ef32769309.fasta |
| Cloned DNA: |
Saccharomyces cerevisiae α-mating factor 1 gene (MFα1, GeneID 855914); prepro secretion signal sequence (ppMF) Trichoderma reesei 1,2-α-D-mannosidase cDNA (mds1); mature sequence |
| Promoter: | Pichia pastoris alcohol oxidase 1 promoter (AOX1) |
| Ribosome binding site: |
- |
| Terminator: | Pichia pastoris alcohol oxidase 1 terminator (AOX1) |
| Selection marker: | Ampicillin (amp) Pichia pastoris HIS4; auxotrophic |
| Replicon: | Escherichia coli plasmid pMB1 origin |
| Host range: | Escherichia coli Pichia pastoris; his4(-), integrative |
| Parental clone: | pPIC9; pGEM11mds1 |
| Further information: | The vector was constructed by inserting a 1.7 kb NarI-NotI fragment of pGEM11mds1, containing the Trichoderma reesei mds1 mature sequence, into the EcoRI-NotI opened pPIC9 vector. The EcoRI and NarI restriction sites were filled in by Klenow DNA polymerase. As a result, the prepro secretion signal sequence was fused to the T. reesei mds1 mature sequence. At the fusion junction, a (Glu-Ala)-dipeptide, as well as additional (but not essential) Tyr-, Val-, Glu-, Phe-, residues are present. During secretion of the protein, the (Glu-Ala)-dipeptide is removed by a dipeptidase. The presence of the P. pastoris HIS4 gene and two P. pastoris AOX1 regions (the AOX1 promoter and the AOX1 3' flanking region) allows integration of the linearized vector into the Pichia genome via homologous recombination at the his4 locus or AOX1 locus respectively. The nucleotide sequence of the T. reesei mds1 cDNA corresponds with the Genbank accession number AF212153.1 except for some amino acid changes described by Maras et al. (2000) and the additional amino acid changes noticed by a BCCM/GeneCorner customer (02/09/02) which have all been confirmed by the sequence analysis results of GeneCorner in 2021. The nucleotide sequence of this plasmid corresponds with the EMBL Nucleotide Sequence Database accession number LT727205.1 except for the latest changes based on the sequence analysis results. Other names of the plasmid are pPICMFmannosidase and pPIC9MFmannase. |
| EMBL Accession number: | AF212153.1, view at EMBL, GenBank, DDBJ LT727205.1, view at EMBL, GenBank, DDBJ |
| Latest sequence update: | 06/10/2021 |
| Sequence detail: | Nucleotide sequence at the fusion junction:
------ ppMF -------> | || || |--- mds1 --->
5' GGG.GTA.TCT.CTC.GAG.AAA.AGA|GAG.GCT||GAA.GCT||TAC.GTA.GAA.TTC.GCC.ACA.AAA
Gly Val Ser Leu Glu Lys Arg|Glu Ala||Glu Ala||Tyr Val Glu Phe Ala Thr Lys
XhoI | || ||SnaBI EcoRI
AvaI HindIII
| : Kex2 cleavage site.
||: dipeptidase cleavage.
Punctuation indicates reading frame.
Overview of the amino acid differences of the mds1 coding sequence:
AA position EMBL Maras2000 Customer SequenceFileGeneCorner
116 S S I I
292 G G A A
298 G G A A
301 G G A A
332 G G A A
361 G G A A
364 T N N N
438 L F F F
496 T N N N |
| Authenticity test: | Restriction enzyme pattern analysed at GeneCorner: BamHI, BglII, EcoRI, FspI and NheI. The abolished BamHI site as a result of the S116I codon change, compared to the EMBL record, was also confirmed by the restriction enzyme analysis results. |
| Class: | Recombinant plasmid |
| Type: | Plasmid |
| History of deposit: | This plasmid was deposited by Prof. Dr R. Contreras(1) (1) Department of Biomedical Molecular Biology, Ghent University, Ghent, Belgium |
| Plasmid reference: | Maras et al., J. Biotechnol. 77 (2000), 255-263 [PMID: 10682284] |
| Related plasmid reference: | Manual of Pichia Expression (Invitrogen) |
| 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: |
pPP1MFmds1 (LMBP 4426) is available at BCCM/GeneCorner. This plasmid was deposited by Prof. Dr R. Contreras Department of Biomedical Molecular Biology, Ghent University, Ghent, Belgium and was published in Maras et al., 2000. |
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.