Arabidopsis Seed Stored mRNAs are Degraded Constantly over Aging Time, as Revealed by New Quantification Methods

How plant seeds age remains poorly understood and effective tools for monitoring seed aging are lacking. Dry seeds contain various stored mRNAs which are believed to be required for protein synthesis during early stages of seed germination. We reasoned that seed stored mRNAs would undergo degradation during seed aging, based on the propensity of mRNAs to degrade. We performed RT-PCR and qPCR analyses to study the changes in stored mRNA levels of Arabidopsis seeds during aging. All stored mRNAs analyzed were gradually degraded in both naturally and artificially aged seeds. The difference in Ct values between aged and control seeds (ΔCt value) was highly correlated with the mRNA fragment size and seed aging time. We derived mathematical equations for estimating the relative amount of undamaged stored mRNAs and frequency of the breakdown at one nucleotide level for individual mRNAs. Stored mRNAs were found to break down randomly. The frequency of breaks per nucleotide per day, which we named β value, remained fairly constant under the same aging conditions over aging time. This parameter should allow the effects of different conditions on the degradation of stored mRNAs to be quantitatively compared. Also, we showed that the change in stored mRNA levels could serve as a more precise biomarker for seed aging assessment than three existing methods. These methods and findings will advance the studies of stored mRNAs and seed ageing in plants, and likely slow RNA degradation in non-plant systems.