Labels 5'-termini prior to partial restriction enzyme digestion.
Unit Definition: One unit is defined as the amount of enzyme required to transfer 1 nmol of g-phosphate from ATP to the 5'-OH termini of salmon sperm DNA fragments in 30 min at 37°C (1).
Storage Conditions: store at -20°C.
Storage Buffer: 50 mM Tris-HCl (pH 7.6 at 22°C), 25 mM KCl, 1 mM dithiothreitol, 0.1 M ATP, 0.1 mM EDTA and 50% (v/v) glycerol.
Assay Conditions: 70 mM Tris-HCl (pH 7.6 at 22°C), 10 mM MgCl2, 5 mM dithiothreitol, 27 nmol of DNA-phosphorus (5'-OH terminated salmon sperm DNA) and 70 nM [ α-³²P]ATP. Reaction volume is 100 µl.
Quality Control: All preparations are tested for contaminating endonuclease, exonuclease and nonspecific RNase activities, along with functional testing in end-labeling reactions.
1.Richardson, C.C. (1971) Progress in Nucleic Acids Research and Molecular Biology 2, 815-828.2. Maxam, A. and Gilbert, W. (1977) Proc. Natl. Acad. Sci. U.S.A. 74, 560-56.
3. Donis-Keller, H. (1980) Nucleic Acids Res. 8, 3133-3142.