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GeneCorner plasmid details

Last data update: 23 October 2020 04:27 CEST

Plasmid name: pCAT1 (LMBP 5032)

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Price category: Cat. 1 (cf. price list)
Status: GeneCorner core plasmid
GeneCorner sequence: p5032.gb (View with Genome Compiler)
p5032.pdf
Sequence
analysis results
Genecorner:

-

Cloned DNA: Nicotiana plumbaginifolia catalase 1 cDNA (CAT1); fragment (CAT1f)
Promoter: Phage T7 gene 10 promoter (T7g10)
Phage T3 promoter
Escherichia coli lac operon promoter
Ribosome
binding site:
-
Terminator: -
Selection marker: Ampicillin (amp)
Replicon: Escherichia coli plasmid pMB1 origin
Phage f1 origin
Host range: Escherichia coli
Parental clone: pBluescriptIIKS+
Further information: The plasmid was constructed by inserting a 1064 bp fragment, containing the 3' region of the Nicotiana plumbaginifolia (tobacco) catalase 1 coding sequence (CAT1), into the EcoRV opened pBluescriptIIKS+ vector.
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 LT727072.1.
The nucleotide sequence of the pBluescriptIIKS+ vector corresponds with the Genbank accession number X52327.1. pBluescriptIIKS+ differs from pBluescriptIIKS- in the orientation of the f1 origin.
The nucleotide sequence of the N. plumbaginifolia CAT1 cDNA corresponds with the Genbank accession number Z36975.1.
EMBL Accession number: Z36975.1, view at EMBL, GenBank, DDBJ
X52327.1, view at EMBL, GenBank, DDBJ
LT727072.1, view at EMBL, GenBank, DDBJ
Latest sequence update: 25/05/2005
Sequence detail:
Nucleotide sequence of the inserted N. plumbaginifolia (tobacco) catalase 1 fragment:

          -- T7 promoter ->  
5' ... GCGTAATACGACTCACTATAGGGCGAATTGGAGCTCCACCGCGGTGGCGGCCGCTCTAGAACTAGTGGATC
                                                     NotI    XbaI  SpeI  BamHI 


                            --------------------- CAT1f --------------------->
                               218                                     492    
      CCCCGGGCTGCAGGAATTCGATGG.AAA.TCA.ACC.TAT.GTG ... GTG.AGA.CCA.AGC.ATA.TGA
       AvaI  PstI  EcoRI    Lys Ser Thr Tyr Val     Val Arg Pro Ser Ile +++
       XmaI                                                          NdeI

    
      ---------- 3'UTR of CAT1 --------->
      AGATGAAGCTTTTAA ... TTTTCTTGTGCTATTATCAAGCTTATCGATACCGTCGACCTCGAGGGGGGGC
           HindIII                       HindIII        SalI  XhoI     ApaI
                                                           HindII

                            <- T3 promoter --
      CCGGTACCCAGCTTTTGTTCCCTTTAGTGAGGGTTAATTGCG ... 3'
        KpnI


+++: Termination codon.
: former EcoRV site.
Punctuation indicates reading frame.
Authenticity test: Restriction enzyme pattern analysed at GeneCorner: AvaI, BamHI, EcoRV, HindIII, KpnI and XmnI.

As compared to the compiled nucleotide sequence, the restriction site analysis pattern revealed that BamHI cuts three times instead of two times, resulting in fragments of approx. 200 bp, 1000 bp and 3000 bp respectively. Consequently, the plasmid seems to be 100 à 150 bp larger than theoretically expected, probably due to a larger 3' UTR part of the insert. The compiled nucleotide sequence has not been adapted accordingly.
Class: Recombinant plasmid
Type: Plasmid
History of deposit: This plasmid was deposited by Prof. Dr D. Inzé(1) (2) and constructed by H. Willekens(1) (2).
(1) Department of Plant Systems Biology, VIB, Ghent, Belgium
(2) Department of Plant Systems Biology, Ghent University, Ghent, Belgium
Plasmid reference: -
Related plasmid reference: Willekens et al., FEBS Lett. 352 (1994), 79-83 [PMID: 7925949]
Restricted distribution: - BCCM MTA
Distributed as: H/P active culture or plasmid DNA
Host for distribution: Escherichia coli K12 XL1-Blue
Host reference: Bullock et al., BioTechniques 5 (1987), 376-379
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:

pCAT1 (LMBP 5032) is available at BCCM/GeneCorner. This plasmid was deposited by Prof. Dr D. Inzé and constructed by H. Willekens .

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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