Dinoflagellate Genomic Organization and Phylogenetic Marker Discovery Utilizing Deep Sequencing Data
Mendez, Gregory Scott
Delwiche, Charles F
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Dinoflagellates possess large genomes in which most genes are present in many copies. This has made studies of their genomic organization and phylogenetics challenging. Recent advances in sequencing technology have made deep sequencing of dinoflagellate transcriptomes feasible. This dissertation investigates the genomic organization of dinoflagellates to better understand the challenges of assembling dinoflagellate transcriptomic and genomic data from short read sequencing methods, and develops new techniques that utilize deep sequencing data to identify orthologous genes across a diverse set of taxa. To better understand the genomic organization of dinoflagellates, a genomic cosmid clone of the tandemly repeated gene Alchohol Dehydrogenase (AHD) was sequenced and analyzed. The organization of this clone was found to be counter to prevailing hypotheses of genomic organization in dinoflagellates. Further, a new non-canonical splicing motif was described that could greatly improve the automated modeling and annotation of genomic data. A custom phylogenetic marker discovery pipeline, incorporating methods that leverage the statistical power of large data sets was written. A case study on Stramenopiles was undertaken to test the utility in resolving relationships between known groups as well as the phylogenetic affinity of seven unknown taxa. The pipeline generated a set of 373 genes useful as phylogenetic markers that successfully resolved relationships among the major groups of Stramenopiles, and placed all unknown taxa on the tree with strong bootstrap support. This pipeline was then used to discover 668 genes useful as phylogenetic markers in dinoflagellates. Phylogenetic analysis of 58 dinoflagellates, using this set of markers, produced a phylogeny with good support of all branches. The Suessiales were found to be sister to the Peridinales. The Prorocentrales formed a monophyletic group with the Dinophysiales that was sister to the Gonyaulacales. The Gymnodinales was found to be paraphyletic, forming three monophyletic groups. While this pipeline was used to find phylogenetic markers, it will likely also be useful for finding orthologs of interest for other purposes, for the discovery of horizontally transferred genes, and for the separation of sequences in metagenomic data sets.