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Misincorporation signatures for detecting modifications in mRNA: Not as simple as it sounds
- Sas-Chen, Aldema, Schwartz, Schraga
- Methods 2019 v.156 pp. 53-59
- RNA-directed DNA polymerase, high-throughput nucleotide sequencing, messenger RNA, transcriptome
- Post-transcriptional modification on mRNA has become a field of intense interest in recent years, and next-generation sequencing based technologies are constantly emerging to detect an increasing number of modifications at a transcriptome-wide level. Some of these approaches are based on identification of misincorporation events induced by reverse transcriptase at modified sites. Although conceptually trivial, sensitive and specific identification of such events is a challenge prone to a surprising number of artifacts, which can lead to substantially inflated estimates of the abundance of diverse modifications. Here we discuss the sources of some of these artifacts and delineate approaches to overcome them.