Tag jumping may produce major distortion of paleoecological reconstructions derived using metabarcoding approaches

Abstract

Analysis of DNA fragments preserved in natural archives, such as lake sediments and buried soil layers, has increased our understanding of past environments and their organism communities.  High throughput sequencing of genetic material from these matrixes and the linkage of DNA barcode databases (e.g. metabarcoding) has played, and still play, an important role for palaeoecological reconstructions. Yet, scrutinization of results derived using metabarcoding approaches – methods that fairly recently have become common within environmental science – is critical. Tag jumping is a process where sample-specific labels (tags), added to DNA sequences to allow identification of individual samples in a pooled DNA library, are unintentionally changed during sampling handling (during the end repair step). If occurring, tag jumping can result in DNA sequences being associated with the wrong sample. Nevertheless, tag jumping is rarely discussed within the paleoecological community, and the importance of the process remains largely unquantified.

Using example datasets from lake sediments and soils, we assessed the impact of tag jumping on metabarcoding data and how it affects paleo-reconstructions. We find that tag jumping can lead to substantial false positive detection of taxa in sedimentary samples. That tag jumping generated a very characteristic topological sample cross-contamination pattern allowed us to estimate that up to 80% of data in some data sets was distorted by this process(Rodriguez‐Martinez et al., 2023). We further compared a set of sedimentary DNA samples processed according to commonly applied, tag-jumping sensitive protocols with the same set of samples processed following a tag jumping free protocol. Our results suggest that tag jumping can affect paleoecological interpretations by: i) masking significant environmental change at a Holocene time-scale by making samples more similar; and; ii) adding species to sediment samples where they were not originally present (false positive detections). Our data clearly shows that tag jumping can represent a major source of bias for paleoecological reconstructions based on metabarcoding techniques. Importantly, the problem is easily circumvented if the right protocols are used.

 

References

Rodriguez‐Martinez, S., Klaminder, J., Morlock, M.A., Dalén, L., Huang, D.Y., 2023. The topological nature of tag jumping in environmental DNA metabarcoding studies. Mol. Ecol. Resour. 1755–0998.13745. https://doi.org/10.1111/1755-0998.13745