Under-representation of short and long 3′ cDNA fragments in 454 sequencing reads. The frequency distribution of 3′ cDNA fragment lengths obtained from in silico digestion of all D. melanogaster transcripts (release 5.1.) is shown in gray. The black line indicates the frequency distribution of 3′ cDNAs obtained from 454 sequencing reads. Independently of the actual counts obtained by the 454 sequencing, each transcript was considered only once. To compare the two datasets that are on different scales, the number of fragments in each class was divided by their root mean square (Becker et al. 1988). After scaling, both samples had a mean of zero and a standard deviation of one. Regardless of which restriction enzyme was used, we noted a pronounced under-representation of short (< ∼80 bp) and long (> ∼300 bp) fragments.
