Previously, cDNAs were constructed to encode various truncated versions of the genome RNA of respiratory syncytial virus (RSV) in which the genome termini and putative transcription signals were retained but the viral genes were deleted and replaced with the reporter gene for bacterial chloramphenicol acetyl transferase (CAT). When introduced into RSV-infected cells, the prototype RSV-CAT minigenome appeared to be replicated, transcribed and packaged into virus-like particles. This represents the first type of system in which genome-like RNAs of a nonsegmented negative strand RNA virus were introduced into the viral replication cycle and rendered biologically active. It is an important step in developing the technology for rescuing complete replication- competent virus from cDNA. This capability would allow the engineering of defined viruses as attenuated vaccines and as reagents for molecular biological and viral pathogenesis studies. In its present form, the RSV- CAT system was used to identify and characterize, for the first time, the cis-acting replication and transcription signals in the RSV genome. It also should be possible to adapt the RSV-CAT system for the analysis of protein function (accompanying report).