 This paper compares five popular transcriptome assembly programs to determine the best approach for generating high-quality assemblies from 454 pyro sequencing data. The authors found that Nubler 2.5 was the best overall performer, producing longer contigs, better alignment to reference sequences, and being faster and easier to use than the other programs. However, they also noted that Nubler 2.3, the current version used by many researchers, did not produce the best results. They recommend combining assemblies from different programs to create a more reliable and accurate final product. This article was authored by Blackster Mark L and Kumar Sujai.