Abstract Background Repeated clusters of ribosomal genes whose activity results in the nucleolus formation are extremely important in multicellular organism genome. Despite the extensive exploration into vertebrate genomes, in many model objects the ribosomal cluster structure is still underinvestigated. So far, complete description for primary structure of avian ribosomal cluster has not been reported. Results This work represents the first successful assembly of complete chicken ribosome cluster sequence. The sequence was deposited to GenBank under accession number KT445934. The total cluster size from pre-rRNA transcriptional start site to the 3′ end of 3’ETS amounted to 11444 bp. 18S rRNA gene size is 1823 base pairs, 5.8S rRNA – 157 bp, 28S rRNA – 4443 bp. The 5 ‘ETS spacer core size is 1839 bp, 3’ETS – about 350 bp., ITS1 – 2099 bp, and ITS2 – 733 bp. The assembly was validated through in situ fluorescent hybridization (FISH) analysis on metaphase chromosomes of chicken. ITS1 and ITS2 spacer sequences have been found to have high GC pair content and form secondary structures featuring high melting temperature. Conclusions Decoding of the chicken rRNA gene cluster sequence extends the use of birds as a model object for exploration into nucleolus organizer region (NOR) regulation and nucleolus functions, e.g. in ontogenesis. This data might also be useful to address certain problems of population and evolutionary genetics.