Investigating low frequency somatic mutations in with Duplex Sequencing.

Mutations are the source of novel genetic diversity but can also lead to disease and maladaptation. The conventional view is that mutations occur randomly with respect to their environment-specific fitness consequences. However, intragenomic mutation rates can vary dramatically due to transcription coupled repair and based on local epigenomic modifications, which are non-uniformly distributed across genomes. One sequence feature associated with decreased mutation is higher expression level, which can vary depending on environmental cues. To understand whether the association between expression level and mutation rate creates a systematic relationship with environment-specific fitness effects, we perturbed expression through a heat treatment in . We quantified gene expression to identify differentially expressed genes, which we then targeted for mutation detection using Duplex Sequencing. This approach provided a highly accurate measurement of the frequency of rare somatic mutations in vegetative plant tissues, which has been a recent source of uncertainty in plant mutation research. We included mutant lines lacking mismatch repair (MMR) and base excision repair (BER) capabilities to understand how repair mechanisms may drive biased mutation accumulation. We found wild type (WT) and BER mutant mutation frequencies to be very low (mean variant frequency 1.8×10 and 2.6×10, respectively), while MMR mutant frequencies were significantly elevated (1.13×10). These results show that somatic variant frequencies are extremely low in WT plants, indicating that larger datasets will be needed to address the fundamental evolutionary question as to whether environmental change leads to gene-specific changes in mutation rate.