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Hidden-break diversity in pancrustacean rRNA profiles

biodiversity
genetics
molecular-biology
zoology

Abstract

Background.
The 28S rRNA hidden break splits the large subunit into two non‑covalently associated fragments (28Sα and 28Sβ), masking 28S in electrophoretic profiles and biasing standard RNA quality metrics in many invertebrates. Pancrustacean diversity in RNA hidden breaks remains incompletely surveyed, particularly for Oligostraca.
Methods.
We sampled 12 species spanning Branchiopoda, Malacostraca, and Oligostraca around Valencia (Spain). RNA was stabilized with DNA/RNA Shield, extracted with Quick‑RNA MagBead, and profiled on an Agilent 5200 Fragment Analyzer. Peaks were assigned to 18S and 28S fragments using BLAST‑inferred gene lengths from reference genomes and annotated rDNA. We analyzed 28S secondary‑structure domains (D‑regions), focusing on D3 and D7a using RNAfold.
Results.
Oligostracans and most branchiopods analyzed showed the canonical single‑peak profile consistent with 18S, 28Sα, and 28Sβ of similar size. Malacostracans exhibited greater profile diversity, including multiple distinct peaks attributable to expansions near D7a that alter 28Sα/β sizes. Comparative analyses indicate conserved D3/D7a architecture across Oligostraca/Branchiopoda and higher variability with frequent expansions in Malacostraca.
Conclusions.
Our data extend RNA profile diversity to Oligostraca, refine fragment‑size estimates with higher‑resolution capillary electrophoresis, and link malacostracan profile heterogeneity to D7a expansions. We recommend rRNA‑aware quality control for arthropod samples and targeted sequencing of poorly sampled lineages (e.g., Mystacocarida, Cephalocarida, Remipedia) to resolve mechanisms and the phylogenetic distribution of the hidden break.
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