Two major RNA species were found in several clonal isolates of avian erythroblastosis virus (AEV) and avian erythroblastosis-associated helper virus (AEAV) complexes: one of 8.7 kilobases (kb), the other of 5.5 kb. The 5.5-kb species was identified as AEV RNA because (i) it was absent from non-transforming AEAV isolated from the same virus complex, (ii) it was present in complexes of AEV and different helper viruses, and (iii) its structure is similar to that of avian acute leukemia viruses of the MC29 group. Molecular hybridization indicated that 54% of AEV RNA is specific and 46% is related to other viruses of the avian tumor virus group, particularly to AEAV, therefore termed group-specific. The genetic structure of AEV RNA was deduced by mapping oligonucleotides representing specific and group-specific sequences and by comparing the resulting map to maps of AEAV and of other avian tumor viruses derived previously. AEV RNA contains a gag gene-related, 5' group-specific section of 1 kb, an internal AEV-specific section of 3 kb unrelated to any other viral RNA tested, and a 3' group-specific section of 1.5 kb. The 5' section of AEV RNA is closely related to analogous 5' sections of the MC29 group viruses and is homologous with a 5' RNA section that is part of the gag gene of AEAV. The 3' section is also shared with AEAV RNA and includes a variant C-oligonucleotide near the 3' end that is different from the highly conserved counterparts of all other exogenous avian tumor viruses. By analogy with Rous sarcoma virus and the acute leukemia viruses of the MC29 group, the internal specific section of AEV RNA is thought to signal a third class of onc genes in avian tumor viruses. Comparisons with AEAV and the MC29 group viruses suggest that both the 5' gag-related and the internal specific RNA sections of AEV are necessary for onc gene function.