Classification performance, at three levels of estimated accuracy (Supplementary Methods), of 6617 possible choices of amplification primer, sequencing primer and read length for single-ended reads from different environments (left portion of each panel) and 3061 possible choices of primer pair and read length for paired-end reads (right portion). Combinations of primers and read lengths are sorted on the x axis according to a measure of overall classification performance (Supplementary Methods). Stacked bars show the proportion of non-chimeric, non-unique sequences from each sample—not the proportion of the total sample—that can be classified to each taxonomic level for each combination. See Supplementary Figure S1 and Supplementary Table S1 for the excluded proportion of novel (and thus a priori unclassifiable) sequences in each sample. The top of each colored section indicates how much of the sample can be classified to the given level or better. ‘Primer miss' (black) indicates sequences that did not match a given primer and so would not be amplified. Classifications more specific than the genus level are exceedingly rare and so are not visible here. Horizontal lines indicate the maximum proportion of each sample classifiable to the genus level using 96 nt or less of sequence (i.e., with an optimal choice of primer or primer pair; see also Supplementary Tables S4 and S5), showing that short reads from the best primers frequently—but not always—provide taxonomic information nearly matching that obtained from longer read lengths. Full-size versions of these panels are available in the supplementary data.