Simple protocol for combined extraction of exocrine secretions and RNA in small arthropods

• Published in: Biology methods and protocols Vol. 9; no. 1; p. bpae054
• Main Authors: Fröhlich, David, Bodner, Michaela, Raspotnig, Günther, Hahn, Christoph
• Format: Journal Article
• Language: English
• Published: England Oxford University Press 2024
• Subjects: biosynthetic pathways; chemical ecology; gland secretion; transcriptomics; chemosystematics; oribatid oil glands; differential expression analysis; phylotranscriptomics
• ISSN: 2396-8923; 2396-8923
• DOI: 10.1093/biomethods/bpae054

The integration of data from multiple sources and analytical techniques to obtain novel insights and answer challenging questions is a hallmark of modern science. In arthropods, exocrine secretions may act as pheromones, defensive substances, antibiotics, as well as surface protectants, and as such they play a crucial role in ecology and evolution. Exocrine chemical compounds are frequently characterized by gas chromatography–mass spectrometry. Technological advances of recent years now allow us to routinely characterize the total gene complement transcribed in a particular biological tissue, often in the context of experimental treatment, via RNAseq. We here introduce a novel methodological approach to successfully characterize exocrine secretions and full transcriptomes of one and the same individual of oribatid mites. We found that chemical extraction prior to RNA extraction had only minor effects on the total RNA integrity. De novo transcriptomes obtained from such combined extractions were of comparable quality to those assembled for samples that were subject to RNA extraction only, indicating that combined chemical/RNA extraction is perfectly suitable for phylotranscriptomic studies. However, in-depth analysis of RNA expression analysis indicates that chemical extraction prior to RNAseq may affect transcript degradation rates, similar to the effects reported in previous studies comparing RNA extraction protocols. With this pilot study, we demonstrate that profiling chemical secretions and RNA expression levels from the same individual is methodologically feasible, paving the way for future research to understand the genes and pathways underlying the syntheses of biogenic chemical compounds. Our approach should be applicable broadly to most arachnids, insects, and other arthropods.