Total tRNA of Chlamydomonas reinhardii was fractionated by 2-dimensional gel electrophoresis. Sixteen tRNAs specific for eleven amino acids could be identified by aminoacylation with Escherichia coli tRNA synthetases. Hybridization of these tRNAs with chloroplast restriction fragments allowed for the localization of the genes of tRNATyr, tRNAPro, tRNAPhe (2 genes), tRNAIle (2 genes) and tRNAHis (2 genes) on the chloroplast genome of C. reinhardii. The genes for tRNAAla (2 genes), tRNAAsn and tRNALeu were mapped by using individual chloroplast tRNAs from higher plants as probes.