Bioluminometric Method for Real-Tim e Detection of Reverse Transcriptase Activity

A simple and sensitive technique for d e tection of reverse transcriptase (RT) activit y in real time has been developed. The tech nique is based on continuous detection o f the inorganic pyrophosphate formed in th e RT-catalyzed reaction by a luminometri c method. The technique has been used fo r continuous monitoring of RT-catalyzed DNA synthesis on both homo- and hetero polymeric templates. The assay is sensitiv e and yields linear responses between 1.5–960 mU of avian myeloblastosis viru s RT (AMV-RT). The assay was used for d e tection of the inhibitory effect o f dideoxythymidine (ddTTP) on the AMV-RT activity and also for real-time detection o f single-base incorporation events catalyzed by AMV-RT. The possibility of using the new technique for other applications is di s cussed .