ABSTRACT
The determination of the sequence of bases in a strand of DNA has developed to the point at which it is an almost trivial operation. Random variations are generally considered minor irritants at best. But in the sequencing of DNA it is crucial that we consider each molecule in a sample as an independent entity which can react in a manner distinct from that of every other molecule. Both “classical” methods, the one developed by Maxam and Gilbert and the other developed by Sanger, rely on electrophoresis of the product mixtures on polyacrylamide gels which are capable of resolving oligonucleotides differing in size by as little as a single nucleotide. In the Maxam–Gilbert method, the DNA sample is first dephosphorylated at both 5’-ends by the action of alkaline phosphatase, and is then rephosphorylated with radioactive phosphate by treating with [32P]-ATP in the presence of polynucleotide kinase.
