A nonacrocentric human chromosome containing transfected human ribosomal DNA was analyzed for participation in nonrandom satellite associations with acrocentric chromosomes. Two constructs were tested. The first contained the complete rDNA transcriptional unit plus flanking DNA and was inserted at chromosome 3p13-->p12 following transfection. The region was silver-positive indicating active transcription. The transcriptionally active rDNA participated in satellite association with other rDNA-bearing chromosomes at a rate 15% above random background associations. The second construct lacked the promoter region. This rDNA integrated at chromosome 4q31-->q32, and was silver stain-negative. Non-random associations with other rDNA-bearing chromosomes were not observed. The study demonstrates that the DNA critical to nonrandom satellite associations is rDNA per se and that active transcription is a requirement.