Seed-stored transcript integrity as a molecular indicator of viability in conserved common bean germplasm

Abstract

Ensuring seed viability over decades is a central challenge in ex situ conservation of plant genetic resources. Standard germination tests, while effective, are destructive and fail to detect early molecular damage that precedes viability loss. RNA integrity has emerged as a promising biomarker due to the inherent vulnerability of mRNA to oxidative degradation in the dry state. In this study, we identified and validated seed-stored mRNAs in common bean (Phaseolus vulgaris), a major crop species extensively represented in global germplasm collections, and investigated transcript degradation patterns in seeds conserved for up to 30 years. Using a comparative genomics approach, we identified 107 P. vulgaris orthologs of long-lived rice mRNAs, many encoding proteins involved in RNA stabilization, oxidoreductase activity, and primary metabolism. Thirty transcripts were validated by RT-qPCR, and the integrity of nine was assessed using paired primers targeting 5′ and 3′ regions. Degradation followed a consistent 5′→3′ pattern, particularly in longer transcripts, and correlated strongly with germination power. Predictive performance varied across transcripts: PSMA4, SMP1, and TRA2 consistently showed strong correlations with viability, whereas others were less informative. The ΔΔCq metric improved resolution by detecting degradation asymmetry. Samples included a genetically diverse panel of conserved, regenerated, and artificially aged accessions, enhancing applicability to real-world genebank conditions. Artificial aging intensified degradation signatures and mirrored patterns observed in naturally aged seeds. Altogether, our results indicate transcript integrity as a molecular biomarker for seed viability, supporting the development of robust molecular tools to inform decision-making and regeneration planning in long-term germplasm conservation.