The autotrophic, sulfur-oxidizing bacterium Thiobacillus denitrificans possesses two forms of the Calvin cycle enzyme ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO). The form I and form II genes were isolated from a cosmid library using heterologous DNA probes. Restriction enzyme analysis indicated that the genes are within 17 kbp of each other. Other Calvin cycle enzyme genes are not present. Analysis of T. denitrificans RNA indicated that the form I genes for the large and small subunits are co-transcribed with a length of 2800 nucleotides. The transcript for the form II gene is 1900 nucleotides in length.