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The Thermo Scientific Biotin DecaLabel DNA Labeling Kit is an advanced system for the efficient synthesis of biotin-labeled DNA probes, based on an improved random-primed labeling method developed by Feinberg and Vogelstein (see References 1, 2). The primary improvement over traditional random-primed method involves the use of random decamers instead of hexamers, ensuring more efficient annealing with DNA at 37°C. Klenow Fragment, exo-, included in the kit, is genetically engineered enzyme with no detectable exonuclease activity. The enzyme does not degrade the labeled probe during reaction, which results in a high labeling yield even with low amounts of template. You can uniformly label any length DNA fragments.
Biotin-labeled DNA is detected with the Biotin Chromogenic Detection Kit or conventional biotin-avidin or biotin-streptavidin detection systems.
Random decamers are annealed to a denatured template DNA molecule, and new strands are synthesized by the Klenow Fragment, exo- in the presence of biotin-dUTP. During this reaction, the biotinylated nucleotides are incorporated into the newly synthesized complementary DNA strand.
DNA labeling by the random primed method. [α-32P]-dNTP, [α-33P]-dNTP, biotin-dUTP, fluorescein-, aminoallyl- or DIG-dUTP can be used.
Dot-blot hybridization with a biotin-labeled probe. Lambda DNA/HindIII was biotin-labeled with the Biotin DecaLabel DNA Labeling Kit and used as a hybridization probe in a dot-blot of the homologous DNA on SensiBlot Plus Nylon Membrane. The blot was developed with the Biotin Chromogenic Detection Kit.
A. P. Feinberg, B. Vogelstein, A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Biochem. 132, 6-13 (1983).
A. P. Feinberg, B. Vogelstein, A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity, Addendum. Biochem. 137, 266-267 (1984).