PAGE 11 OF 16 D Nucleic Acids Research, 2005, Vol. 33, No. 13 e115 Figure 7. Distribution of codons in different sections of genes. The figures (A–D) are for valid oligos in genome data. were calculated based on the nucleotide content. The codons were analyzed according to the gene, i.e. the coding region within oligos started either from the ﬁrst, second or third position dependent on the match with the gene sequence. In prokaryotes, the distributions are rather normal for all the codons except for in C.acetobutylicum, S.solfataricus and M.tuberculosis, which show large deviations for most of the codons. Even higher deviations are apparent in the eukaryal genomes. It is intriguing, that in most instances all the eukaryotes have similar trends for a large number of codons although the extent of the bias varies. This is of notion because these organisms have different codon preferences. We compared further the Z-values for all codons (Figure 7A–D). Synonymous codons are known to have strong bias. Codon usage has effect, for example, on the translation. Highly expressed genes contain mainly those codons for which there are abundant tRNAs. The codon usage varies between organisms. There were no general trends for the usage of codons. When looking at the codon usage within the seven sections of genes, the majority of the codons in the majority of organ- isms have a normal distribution. The distribution is almost equal for the majority of codons in each section. However, certain patterns are visible. The largest, eukaryal genomes have the highest Z-scores, especially A.thaliana, which had signi- ﬁcant bias in many places. Also C.elegans and S.cerevisiae have biased distribution to sections, but not that often and generally the Z-values are smaller than for A.thaliana, which is clearly the largest of the studied genomes. Clear examples of C+G-rich codons for alanine and glycine are A.tumefaciens and M.tuberculosis, which have signiﬁcantly less codons con- taining C or G in the third position, although these organisms have the highest overall C+G content. Also S.typhimurium is biased towards not having G at the third position in codon for alanine. C.acetobutylicum has strong bias in a number of codons, and S.solfataricus and A.fulgidus in some individual cases. The genomes of prokaryotes have less biased distribu- tion. Usually, the eukaryotes clearly favor certain triplets when synonymous codons appear.