pSCH2003 plasmid in E. coli (ATCC® 87427)

Organism: Unidentified bacterium  /  Clone Type: Clone  /  Depositors: KJ Shaw

Permits and Restrictions

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Designations pSCH2003 plasmid in E. coli
GenBank Number

X04555

Species Unidentified bacterium
Depositors KJ Shaw
Vector
Construct size (kb): 3.25
Insert
DNA: Synthetic
Insert lengths(kb): 0.305
Gene product: aminoglycoside adenylyltransferase(spectinomycin resistance, streptomycin resistance) [ant(2")-Ia]
Insert Size (kb) 0.305
Media ATCC® Medium 1227: LB Medium (ATCC medium 1065) with 50 mcg/ml ampicillin
Biosafety Level 1

Biosafety classification is based on U.S. Public Health Service Guidelines, it is the responsibility of the customer to ensure that their facilities comply with biosafety regulations for their own country.

Comments
Restriction digests of the clone give the following sizes (kb): XhoI--3.3; HindIII--3.3; PstI--3.3; HindIII/XhoI--2.8, 0.4.
The insert contains the following restriction sites (approximate kb from the 5' end): PvuII--0.20; RsaI--0.25; SphI--0.29.
A single gel purification of the PCR generated probe is necessary since flanking regions will co-amplify with the gene specific sequence. Failure to do so often results in high backgrounds and false positives with clinical E. coli strains.
A hybridization probe may be generated using the following vector specific PCR primers: modified T3 = 5'-CCCCTCACTAAAGGGAACAAAAGCTG-3' and modified T7 = 5'-CGCGTAATACGACTCACTATAGGGCGAA-3'.
References

Shaw KJ, et alThe application of molecular techniques for the study of aminoglycoside resistanceIn: Shaw KJ, et alMethods in molecular medicine: molecular approaches for the diagnosis and investigation of bacterial diseasesTotowa, NJHumana Presssubmitted, 1996

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