Degraded RNA profiles in Arthropoda and beyond

The requirement for high quality non-degraded RNA is essential for an array of molecular biology analyses. When analysing the integrity of rRNA from the barnacle Lepas anatifera (Phylum Arthropoda, Subphylum Crustacea), atypical or suboptimal rRNA profiles that were apparently degraded were observed on a bioanalyser electropherogram. It was subsequently discovered that the rRNA was not degraded, but arose due to a gap deletion (also referred to as hidden break) in the 28S rRNA. An apparent excision at this site caused the 28S rRNA to fragment under heatdenaturing conditions and migrate along with the 18S rRNA, superficially presenting a degraded appearance. Examination of the literature showed similar observations in a small number of older studies in insects; however, reading across multiple disciplines suggests that this is a wider issue that occurs across the Animalia and beyond. The current study shows that the 28S rRNA anomaly goes far beyond insects within the Arthropoda and is widespread within this phylum. We confirm that the anomaly is associated with thermal conversion because gap-deletion patterns were observed in heat-denatured samples but not in gels with formaldehyde-denaturing.