FIGURE 7.

The induction of RCA by hybridization of the 3′ end of target RNA. The experiments (A) were performed under the conditions described in “Materials and Methods.” The 5′-end-labeled RNA1–DNA hybrids were incubated with Phi29 DNA polymerase exo− mutant (D12A/D66A) at 37°C in the presence or absence of 1 mM dNTPs. After incubation the samples were divided into two equal parts, one of which was treated with the excess of DNaseI. The reaction products were analyzed by electrophoresis through denaturing 8% polyacrylamide gels. RNA1 substrate is shown above the gels. The radioactive label is indicated by the star. The reaction scheme (B) represents the conversions of target RNA. Reaction substrate and products are labeled as “a” for the 5′-end-labeled RNA1 target serving as an RNA primer for RCA, “b” for the 5′-endlabeled RCA product, and “c” for the 5′-end-labeled RCA product after DNaseI treatment comprising the RNA target with residual DNA.