Labeling of Double-Stranded DNA Probes with Biotin

Probes labeled with biotin have been used for hybridizations to Southern blots and to chromosomes in situ since the early 1980s (1 ). Nowadays, the labeling of double-stranded DNA probes with biotin is made easy by the availability of labeling kits that provide everything required for nick translation of DNA probes in the presence of biotinylated UTP. The method described here is based on the use of the BRL (Bethesda Research Laboratories, Gaithersburg, MD) BIONICK Labeling Kit (catalog no. 8247SA). While the labeling reaction is taking place, a fractionation column is prepared to purify the biotin-labeled DNA. After the fractions have been collected, incorporation is assessed on test strips using a BRL detection kit (catalog no. 8239SA). The whole procedure takes a whole day, with a convenient break when the test strips are incubating in the vacuum oven (see step 18 below). Several probes may be labeled in one experiment, although four probes is about the optimum number to handle by a single worker.