A kinetic equation was derived for complex DNA polymerase reactions. The complex repertoire of polymerase reactions includes the facts that the four monomeric substrates involved in the polymerase reactions may also work as competitive inhibitors depending on their incorporation sites, the incorporation rates of each deoxynucleotide are different, and polymerization proceeds in parallel at various sites on each template DNA. Using the equation, the kinetic properties of Thermus thermophilus DNA polymerase were investigated. The reaction at the A-incorporation site is strongly inhibited by dTTP and dGTP with inhibition constants of 9.9 and 7.1 mu M, respectively. The reaction at the T-incorporation site is also inhibited by dATP with an inhibition constant of 7.6 mu M, while the G- and C-incorporation sites are less sensitive to competitive inhibition. When substrate concentrations are low, the incorporation rate of A is the fastest and that of C is the slowest. Accordingly, the C-incorporation step largely limits the overall reaction rate. Using the kinetic parameters determined experimentally, the optimum ratio of the concentrations of the four nucleotides was calculated.