Application of next generation long read sequencing to protistan systematics

Abstract

Life appeared on this planet around 3.5 billion years ago (bya), as unicellular organisms without nuclei. Current phylogenetic analysis divides life in three domains: Bacteria, Archaea and Eukarya. In order to understand evolutionary events in the past, it is crucial that phylogenetic information is correlated to the fossil record. Unfortunately, the vast majority of microbial diversity does not leave a reliable fossil record. One fortuitous exception are the Arcellinida, a lineage of amoeboid organisms that construct an outer carapace (popularly known as test). The Arcellinida fossil record has recently been corroborated to relate Tonian Vase-Shaped Microfossils, dating around 750 million years ago. This discovery has enabled deeper understanding of the climatic events that lead to the deep ocean oxygenation that occurred in the Neoproterozoic. Although reliable, the current phylogenomic tree of Arcellinida still lacks taxonomic sampling. Here, we propose to expand sampling by including two enigmatic genera: Schoenbornia and Microcorycia. Each genus is critical for a different reason: Schoenbornia has intermediate morphological characters that can clarify events in the evolution of agglutinated shells, and Microcorycia is theoretically the type species of the recently described group Corycida, which is group of shelled amoebae that was recently shown to group outside of Arcellinida. We have been able to identify these rare organisms in recent samples in the laboratory. We propose to use the already established technique of Single-Cell transcriptomic sequencing using Illumina technology to generate the phylogenomic dataset. This technique is routine in our laboratory, but we will also do a comparison with the newer generation of long-read sequencing provided by Nanopore's MinION technology, which will advance the laboratory into the leading edge in sequencing and generate expertise that is currently lacking. (AU)