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Abstract for Terminal Deoxynucleotidyl Transferase
Terminal Deoxynucleotidyl Transferase (TdT) is an enzyme that catalyzes the repetitive addition of mononucleotides from dNTPs to the terminal 3´-OH of a DNA initiator, accompanied by the release of inorganic phosphate. The enzyme thus provides a unique method for the labeling of the 3´ termini of DNA.
This technical bulletin provides two different protocols for the incorporation of 32P. The first illustrates the addition of an [alpha-32P]dNTP “tail” to the 3´ termini of single-stranded DNA fragments. In addition, incorporation can be limited to a single nucleotide by using [alpha-32P]cordycepin-5´-
triphosphate. This analog lacks a free 3´ hydroxyl group, preventing incorporation of additional nucleotides.