The amount of DNA synthesis in vitro with the ultraviolet-irradiated poly-(dT) . oligo(rA) template initiators catalysed by DNA polymerase alpha (Masaki, S. and Yoshida, S., Biochim. Biophys. Acta 521, 74--88) decreased with the dose of ultraviolet-irradiation. The ultraviolet irradiation to the template, however did not affect the rate of incorporation of incorrect deoxynucleotides into the newly synthesized poly(dA). The addition of terminal deoxynucleotidyl transferase to this system enhanced the DNA synthesis to a level which is comparable to that of the control and it concomitantly increased the incorporation of the mismatched deoxynucleotide into the newly synthetized poly(dA) strands. On the other hand, with an unirradiated template initiator, the misincorporation was only slightly enhanced by the addition of terminal deoxynucleotidyl transferase. The sizes of newly synthetized DNA measured by sedimentation velocities were found to be smaller with the ultraviolet-irradiated templates but they increased to the control level with the addition of terminal deoxynucleotidyl transferase to the systems. These results suggest that terminal deoxynucleotidyl transferase can help DNA polymerase alpha to "bypass" thymine dimers in vitro by the formation of mismatched regions at the positions opposite to pyrimidine dimers on the template.