Complementary DNA (cDNA) clones specific to the smallest chromosome (chromosome I) of Chlorella vulgaris C-169 were selected from cDNA libraries with probes of chromosome I DNA fragments amplified by degenerate oligonucleotide-primed polymerase chain reaction (DOP-PCR). A total of 15 clones was obtained, which included gene homologs for alpha -tubulin, inosine-5'-monophosphate dehydrogenase, beta -1,4-mannase, a TTG-binding protein, a heat shock protein, thioredoxin/protein disulfide isomerase, transcription factor NF-E2, an oxidoreductase, and UDP-n-acetylglucosamine enolpyruvyltransferase. These clones were definitely localized at specific Sites on the chromosome I physical map constructed on the basis of overlapping cosmid clones (the contig). They were predominantly distributed within the left two-thirds of the chromosome. This contrasts with the distribution of repetitive:elements such as short interspersed elements (SINEs), which are rather abundant in the right two-thirds of chromosome I. The comparative simplicity of the gene arrangement of Chlorella chromosome I suggests that it may be able to serve as a prototypic system for deciphering the complexity of huge plant chromosomes.