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Identification of Homologous Recombination Events in Mouse Embryonic Stem Cells Using Southern Blotting and Polymerase Chain Reaction
Published on: November 20, 2018
Detection of specific DNA sequences-the southern transfer
1MRC Molecular and Cellular Cardiology Research Unit, University of Stellenbosch Medical School, Tygerberg, South Africa.
Abstract:
The purpose of this technique is the detection and characterization of specific DNA sequences. The DNA is fragmented by digestion with a restriction endonuclease, and the fragments separated by agarose gel electrophoresis. The DNA is then denatured in the gel and transferred to a nitrocellulose filter. This is incubated with a (32)P-labeled probe, which is DNA having a base sequence complementary to the DNA that is to be detected on the filter. After hybridization of the probe to its complementary sequence, unbound probe is washed off. The position of the probe on the filter is then detected by autoradiography. This procedure was developed by E. M. Southern of Edinburgh University (1), and is generally referred to as the Southern transfer or Southern blot.
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