In vitro synthesis of short DNA pieces by DNA polymerase alpha from mouse myeloma.


The DNA chain elongation mechanisms of mouse DNA polymerases alpha and ...
The DNA chain elongation mechanisms of mouse DNA polymerases alpha and beta have been analyzed by using denatured DNA with a (dT)n block at the 3'-end as a template in combination with RNA ((rA)12-20)primer. The (rA)12-20-primed DNA product synthesized by DNA polymerase alpha was 3-5 s in size even after prolonged reaction; instead of a size increase, the number of 3-5 s molecules increased with the reaction time. The size of products was not affected by differences in 3H-labeled substrate (dATP or dTTP), enzyme amount, KCl concentration, or the length of 3'-(dT)n blocks. On the other hand, DNA polymerase beta synthesized long DNA products by a highly distributive reaction mechanism. 3-5 sDNA pieces synthesized by DNA polymerase alpha were not elongated any further by DNA polymerase alpha, but were converted into long DNA chains by DNA polymerase beta. The results imply that DNA polymerase alpha recognizes the size of the product DNA, and shuts off further elongation.




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