Sequence comparison of the chloroplast genes of the large subunit of ribulosebisphosphate carboxylase from wild-type and from a uniparental mutant of the green unicellular alga Chlamydomonas reinhardii has revealed a single nucleotide change. The corresponding Gly to Asp amino acid substitution would introduce a negative charge into the presumptive substrate binding region of the enzyme and would explain the inactivity of the mutant protein. This is the first chloroplast mutation whose DNA sequence is known. Our results establish the first exact point of correlation between the physical map of the chloroplast genome of C. reinhardii and a specific genetic locus.