We have previously described the experimental introduction of recombinant subgroup A avian leukosis viruses (ALV) with Rous-associated virus 0 long terminal repeats into the germ line of line 0 chickens and the generation of 23 transgenic lines. Two of these transgenic lines, alv6 and alv11, do not produce infectious virus. Both of these lines contain defective proviruses but do express the gag and/or env protein. We have measured viral RNA expression in tissues derived from alv6, alv11, and the parental line 0. Total RNA was prepared from 9-day embryo, 16-day embryo, 1-day chicken, and 28-day chicken tissues. Viral RNA was detected by Northern RNA transfer analysis. The results indicate that both alv6 and alv11 chickens express viral RNA in all tissues tested regardless of the stage of development. No viral transcripts were detected in any line 0 (C/E; ev-negative) tissue. The levels of biologically active env glycoprotein correlates with the env RNA levels in both lines. In an in vivo interference assay, alv6, alv11, and line 0 chickens were infected with Rous-associated virus 1 and monitored for viremia, antibody against Rous-associated virus 1, and ALV-induced pathogenesis from 4 to 21 weeks. None of the 61 alv6 chickens contained detectable virus or produced antibody against subgroup A ALV. Virus and/or antibody against subgroup A ALV was detected in 34 of the 43 alv11 chickens, whereas 51 of 52 line 0 birds were viremic and/or produced antibody. ALV-induced pathogenesis was observed predominantly in line 0 chickens (10 of 59), whereas very little ALV-induced pathogenesis was seen in either alv6 (1 of 62) or alv11 (1 of 44) chickens. Presumably the mechanism for the increased resistance of alv6 and alv11 chickens was subgroup-specific receptor interference. These results clearly demonstrate that experimentally introduced endogenous proviruses can be expressed at high levels in the avian system.