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GeneCorner plasmid details

Last data update: 11 June 2021 14:12 CEST

Plasmid name: pUChGNTI1 (LMBP 3133)

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Price category: Cat. 1 (cf. price list)
Status: GeneCorner core plasmid
GeneCorner sequence: p3133.gb (View with Genome Compiler)
p3133.pdf
Sequence
analysis results
Genecorner:

-

Cloned DNA: Human N-acetylglucosaminyltransferase I cDNA (GlcNAc-TI, MGAT1)
Promoter: Escherichia coli lac operon promoter
Ribosome
binding site:
Ribosome binding site (RBS) of the Escherichia coli lac Z gene (lacZ)
Terminator: -
Selection marker: Ampicillin (amp)
Replicon: Escherichia coli plasmid pMB1 origin
Host range: Escherichia coli
Parental clone: pUC18
Further information: The plasmid was constructed as follows: 1) the human N-acetylglucosaminyltransferase I cDNA (hGlcNAc-TI) was obtained after PCR on colon carcinoma gDNA that contains no introns; 2) the obtained PCR fragment, carrying a 3' sticky 'A' nucleotide due to the use of Pfu DNA polymerase for the PCR procedure, was cloned into pUC18, previously digested with HindII and tailed with dideoxythymidine triphosphate using terminal transferase according to Holton et al. (1991).
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 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: 15/12/1994
Sequence detail:
Nucleotide sequence at the start of the human GlcNAc-TI coding sequence:

   pUC18 -->   --> PCR fragment
5' TCTAGAGTC T CCAGG ATG.CTG.AAG. 3'
   XbaI  --- ^        *
         blunt-ended HindII

^: T-residue, due to tailing.
*: Start codon of the human GlcNAc-TI coding sequence.


Nucleotide sequence at the end of the human GlcNAc-TI coding sequence:

             PCR fragment -->   --> pUC18
5' TGG.AAT.TAG CACCTGTCGACTGG A GACCTG 3'
            *       HindII    ^ ---
                    SalI        blunt-ended HindII
                    AccI

^: 3' sticky A-residue, due to the PCR procedure.
*: Termination codon of the human GlcNAc-TI coding sequence.
Authenticity test: Restriction enzyme pattern analysed at GeneCorner: BglI, Eco47III, HindII and NarI/XmnI.
Class: Recombinant plasmid
Type: Plasmid
History of deposit: This plasmid was deposited by Dr M. Maras(1)(2) and Prof. Dr R. Contreras(1)(2).
(1) Department for Molecular Biomedical Research, VIB, Ghent, Belgium
(2) Department of Biomedical Molecular Biology, Ghent University, Ghent, Belgium
Plasmid reference: Maras et al., Eur. J. Biochem. 249 (1997), 701-707 [PMID: 9395316]
Related plasmid reference: Maras et al., FEBS Lett. 452 (1999), 365-370 [PMID: 10386623]
Holton et al., Nucleic Acids Res. 19 (1991), 1156 [PMID: 2020554]
Kumar et al., Proc. Natl. Acad. Sci. U.S.A. 87 (1990), 9948-9952 [PMID: 1702225]
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:

pUChGNTI1 (LMBP 3133) is available at BCCM/GeneCorner. This plasmid was deposited by Dr M. Maras and Prof. Dr R. Contreras and was published in Maras et al., 1997.

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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