Practice question

Five-prime end labeling with radioactive phosphate relies on the forward reaction catalyzed by T4 polynucleotide kinase. The enzyme transfers the terminal phosphate from ATP to the free 5'-hydroxyl of dephosphorylated DNA or RNA, forming a new phosphomonoester. When gamma-32P-ATP of high specific activity is supplied, the labeled phosphate is incorporated directly at termini, generating probes suitable for autoradiographic detection, DNA footprinting, and restriction mapping. Prior dephosphorylation of substrate with alkaline phosphatase dramatically enhances incorporation by increasing available hydroxyl acceptors and preventing exchange reaction limitations.