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LMBP plasmid details

Last data update: 05 March 2021 04:23 CET

Plasmid name: YDp-L (LMBP 3355)

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Price category: Cat. 1 (cf. price list)
Status: GeneCorner core plasmid
GeneCorner sequence: p3355.gb (View with Genome Compiler)
p3355.txt
p3355.pdf
Sequence
analysis results
Genecorner:

-

Cloned DNA: Saccharomyces cerevisiae leucine 2 disruption cassette (LEU2)
Promoter: Escherichia coli lac operon promoter
Ribosome
binding site:
-
Terminator: -
Selection marker: Ampicillin (amp)
LEU2; auxotrophic
Replicon: Escherichia coli plasmid pMB1 origin
Host range: Escherichia coli
Saccharomyces cerevisiae; leu2(-), integrative
Parental clone: pUC9HStop; pJH-L1
Further information: The plasmid was constructed by inserting a 1601 bp (instead of 1613 bp as mentioned in Berben et al. (1991); personal communication Dr G. Berben) HpaI-AccI (filled in with Klenow DNA polymerase) fragment of pJH-L1, containing the S. cerevisiae LEU2 auxotrophically selectable marker gene, into the unique HindIII site (filled in with Klenow DNA polymerase) of pUC9HStop, a pUC9 derivative differing in the sequence of the multicloning site.
This is an integrative yeast plasmid designed for gene disruption purposes. The disruption cassette consists of the LEU2 gene, flanked by termination codons in all three reading frames and restriction sites. The termination codons prevent read-through from the remaining sequences of disrupted genes and those of the selectable disrupting fragment; in this way, possible artefactual phenotypic effects caused by fortuitous hybrid proteins are avoided.
When cloning a fragment downstream from the lac promoter it may be advisable to use lacI(q) strains in order to prevent fortuitous expression of a possibly noxious polypeptide.
The nucleotide sequence of this plasmid corresponds with the EMBL Nucleotide Sequence Database accession number LT727471.1.
The nucleotide sequences at the junctions were experimentally verified by Dr G. Berben prior to deposit at BCCM/GeneCorner.
EMBL Accession number: LT727471.1, view at EMBL, GenBank, DDBJ
Latest sequence update: 12/10/1995
Sequence detail:
Nucleotide sequence at the termination modules flanking the LEU2 insert:

                                  HindIII/HpaI fusion 
                     stop module       |
                     ---------->       |--> LEU2 DNA
5' GAATTCCCGGGGATCCGGTGATTGATTGAGCAAGCT|AACTGTGG ...
   EcoRI     BamHI   --- --- ---  
        SmaI



        AccI/HindIII fusion
            |       stop module
--> LEU2 DNA|       <----------
... TAGGGTAG|AGCTTGCTCAATCAATCACCGGATCCGTCGACCTGCAGCCAAGCTAGCTT 3'
                    --- --- ---  BamHI SalI  PstI      NheI
Authenticity test: Restriction enzyme pattern analysed at GeneCorner: AccI/NarI, BamHI/HindIII, EcoRI/PstI, HaeII/HpaI, SmaI and XmnI.
Class: Recombinant plasmid
Type: Plasmid
History of deposit: This plasmid was deposited by Dr G. Berben(1).
(1) Centre de Recherches agronomiques de l' Etat, Station de Chimie et de Physique agricoles, Laboratoire de Microbiologie, Gembloux, Belgium
Plasmid reference: Berben et al., Yeast 7 (1991), 475-477 [PMID: 1897313]
Related plasmid reference: Bax et al., RNA 12 (2006), 2005-2013 [PMID: 17018574] [DOI: 10.1261/rna.159406]
Restricted distribution: - BCCM MTA
Distributed as: H/P active culture or plasmid DNA
Host for distribution: Escherichia coli K12xB HB101
Host reference: Boyer and Roulland-Dussoix, J. Mol. Biol. 41 (1969), 459-472 [PMID: 4896022]
Related host reference: Sambrook et al. (eds), Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory Press, NY (1989) [ISSN/ISBN: 0879693096]
Cultivation medium: LB-Lennox + ampicillin (50 μg/ml)
Cultivation temperature: 37°C
Biosafety level: L1
Cultivation remark: Cultivation conditions: preferably under rotational shaking.
Other culture collection numbers: -

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.

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