The timing of methylation of the ribosomal sequences of ribosomal precursor RNA (pre-rRNA) from the yeast Saccharomyces carlsbergensis was investigated by fingerprint analysis of the methylated oligonucleotides derived from the various precursors. From the total of 37 ribose and 6 base-methyl groups found in 26-S rRNA, the two copies of the base-methylated nucleoside m3U as well as the doubly methylated sequence Um-Gm psi are not yet present in 37-S RNA, the predominant common precursor of 26-S and 17-S rRNA. Introduction of these methyl groups into the ribosomal sequences appears to take place at the level of 29-S pre-rRNA, the immediate precursor to 26-S rRNA. From the total of 18 ribose-methylated and 6 base-methylated nucleosides found in 17-S rRNA, the latter group (one copy of m7G, the m62A-m62A- sequence and the hypermodified methylated nucleoside "mX") is completely missing in 37-S pre-rRNA. The methyl group of m7G is introduced into 18-S pre-rRNA, the direct precursor of 17-S rRNA, in the nucleus. The -m62A-m62A- sequence is methylated after transport of the 18-S pre-rRNA to the cytoplasm prior to the final maturation into 17-S rRNA.