Rapid one-step characterization of recombinant vectors by direct analysis of transformed Escherichia coli colonies.
We have developed a polymerase chain reaction (PCR) based procedure for rapidly analyzing recombinant vectors in whole bacterial cells. No purification, restriction mapping or sequencing of vectors is required and the results are available within 6 hours. Whole cells are added to a PCR mix that is designed to amplify DNA only if the correct insert is present in the required orientation. The presence of an appropriately sized band on an agarose gel is indicative of a correct clone.
Sandhu
G S
GS
Dept. of Biochemistry and Molecular Biology, Mayo Graduate School of Medicine, Mayo Clinic and Foundation, Rochester, MN 55905.
Precup
J W
JW
Kline
B C
BC
D001709:Biotechnology; D004247:DNA / Q000235:genetics; D004269:DNA, Bacterial / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics*; D005169:Factor VIII / Q000235:genetics; D005822:Genetic Vectors; D006801:Humans; D021141:Nucleic Acid Amplification Techniques; D016133:Polymerase Chain Reaction / Q000379:methods*; D011995:Recombination, Genetic; D014170:Transformation, Genetic
D004269:DNA, Bacterial; D005169:Factor VIII; D004247:DNA