UNIVERSITY OF CALIFORNIA, SANTA CRUZ MAMIELLOPHYCEAE: PHYLOGENETIC AND BIOGEOGRAPHIC INSIGHTS A dissertation submitted in partial satisfaction of the requirements for the degree of DOCTOR OF PHILOSOPHY in OCEAN SCIENCE by Melinda P. Simmons June 2014 The Dissertation of Melinda Simmons is approved by: _________________________________ Associate Adjunct Professor Alexandra Z. Worden, Chair _________________________________ Professor Jonathan P. Zehr _________________________________ Associate Adjunct Professor Steven H. D. Haddock _________________________________ Professor Raphael Kudela ___________________________________ Tyrus Miller Vice Provost and Dean of Graduate Studies TABLE OF CONTENTS Title Page………..………………………...……………………………………….…. i Table of Contents……………..……………………………………………………... iii List of Tables and Figures……..………………… ………….………..……………. vi Abstract……………………………..…….……….……………………………...... viii Acknowledgments…….……………………………..………….…………….…........ x 1 Chapter 1: Introduction .......................................................................................... 1 1.1 The Importance of Marine Phytoplankton within the Class Mamiellophyceae ...................................................................................................... 1 1.2 Diversity within the Mamiellophyceae .......................................................... 2 1.3 Mamiellophyceae Abundance ........................................................................ 5 1.4 Biogeography of Mamiellophyceae ............................................................... 8 1.5 Aims and Objectives .................................................................................... 10 2 Chapter 2: Micromonas Genome Architecture .................................................... 16 2.1 Abstract ........................................................................................................ 16 2.2 Introduction .................................................................................................. 17 2.3 Methods ........................................................................................................ 20 2.3.1 Randomized analysis of gene structure .................................................... 20 2.3.2 Introner element analysis .......................................................................... 21 2.4 Results .......................................................................................................... 21 2.5 Discussion .................................................................................................... 26 iii 2.5.1 COP Formation ......................................................................................... 26 2.5.2 COP Function ........................................................................................... 27 2.5.3 Introner Elements ..................................................................................... 31 2.6 Conclusion .................................................................................................... 32 3 Chapter 3: Prolific but distinct repetitive introns in picoeukaryote species highlight Antarctic Micromonas ................................................................................. 47 3.1 Abstract ........................................................................................................ 47 3.2 Introduction .................................................................................................. 48 3.3 Methods ........................................................................................................ 52 3.3.1 Culturing and nucleic acid extraction ....................................................... 52 3.3.2 PCR, cloning and sequencing ................................................................... 54 3.3.3 Clustering and phylogenetics .................................................................... 55 3.3.4 Metagenome searches ............................................................................... 57 3.4 Results .......................................................................................................... 58 3.5 Discussion .................................................................................................... 68 3.6 Conclusions .................................................................................................. 75 4 Chapter 4: Biogeography of photosynthetic picoeukaryotes in the North Pacific Ocean .......................................................................................................................... 90 4.1 Abstract ........................................................................................................ 90 4.2 Introduction .................................................................................................. 91 4.3 Materials and Methods ................................................................................. 93 iv 4.3.1 Field sampling .......................................................................................... 93 4.3.2 Flow cytometry ......................................................................................... 94 4.3.3 QPCR ........................................................................................................ 95 4.3.4 Metatranscriptome library construction and sequencing .......................... 96 4.3.5 Metatranscriptome analyses ..................................................................... 98 4.3.6 Determining genetic divergence among putative Bathycoccus and Ostreococcus ecotypes ....................................................................................... 100 4.3.7 Determining Ecomarkers ........................................................................ 101 4.3.8 Identifying Ecotype Ecomarkers in Meta-omic Data ............................. 102 4.3.9 Analysis of SOD and nitrogen transporter gene families ....................... 102 4.4 Results & Discussion ................................................................................. 103 4.4.1 Environmental conditions ....................................................................... 103 4.4.2 Phytoplankton abundance ....................................................................... 104 4.4.3 Establishment of Bathycoccus ecotype genetic distances ...................... 109 4.4.4 Diversity assessment based on metatranscriptomic data ........................ 110 4.4.5 Exploratory gene expression analyses .................................................... 113 4.5 Conclusion .................................................................................................. 118 5 Chapter 5: Conclusions and Perspectives .......................................................... 137 v List of Tables and Figures Table 1.1 ..................................................................................................................... 12 Table 2.1 ..................................................................................................................... 34 Table 2.2 ..................................................................................................................... 34 Table 2.3 ..................................................................................................................... 35 Table 2.4 ..................................................................................................................... 35 Table 2.5 ..................................................................................................................... 36 Figure 2.1 .................................................................................................................... 37 Figure 2.2 .................................................................................................................... 38 Figure 2.3 .................................................................................................................... 39 Figure 2.4 .................................................................................................................... 39 Figure 2.5 .................................................................................................................... 40 Figure 2.6 .................................................................................................................... 41 Table 3.1. .................................................................................................................... 77 Table 3.2. .................................................................................................................... 78 Table 3.3. .................................................................................................................... 79 Figure 3.1 .................................................................................................................... 80 Figure 3.2 .................................................................................................................... 81 Figure 3.3 ................................................................................................................... 82 Table 4.1 ................................................................................................................... 119 Table 4.2 ................................................................................................................... 120 Table 4.3 ................................................................................................................... 122 Figure 4.1 .................................................................................................................