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Identification, Characterization and Evolution of the Mating Type Locus in Diatoms

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Identification, Characterization and Evolution of the Mating Type Locus in Diatoms Department of Biology Department of Plant Biotechnology and Bioinformatics Department of Plant Systems Biology Identification, characterization and evolution of the mating type locus in diatoms Ives Vanstechelman Promotors: Prof. Dr. Koen Sabbe and Prof. Dr. ir. Marnik Vuylsteke Thesis submitted in fulfillment of the requirements for the degree of Doctor (PhD) in Sciences (Biology) Academic year 2012-2013 Exam commission Promotors: Prof. Dr. Koen Sabbe Prof. Dr. ir. Marnik Vuylsteke Members of the reading commission Dr. Mariella Ferrante Prof. Dr. Olivier De Clerck Prof. Dr. Wout Boerjan Other members of the exam commission Prof. Dr. Wim Vyverman Prof. Dr. Mieke Verbeken Dr. Marie Huysman Copyright © Ives Vanstechelman, Department of Biology, Faculty of Sciences, Ghent University, 2013. All rights reserved. No part of this publication may be reproduced, stored in a retrieval system, or transmitted, in any form or by any means, electronic, mechanical, photocopying, recording, or otherwise, without permission in writing from the copyright holder(s). Acknowledgements The work presented in this thesis could not be successful without the help of several people. Therefore, I want to give a few words of thanks and appreciation for the people who supported me during this 4 years during PhD. First of all, I would like to thank my two promotors Koen Sabbe and Marnik Vuylsteke. Koen is my promotor with the biological background. He supported me a lot in giving more knowledge about the biology of diatoms and evolution. When I first met him, he could directly convince me to start in this position. The unique features of the diatom life cycle attracted me and I really saw it as a big challenge to get insight into the genetic basis of this. The person who helped me the most with succeeding in this PhD is undoubtedly Marnik Vuylsteke. He is my promotor with the genetic background and as a consequence, I mainly worked in his lab, the Quantitative Genomics lab. Making progression in a PhD project is the same as being constanly confronted with a lot of problems and I am sure, Marnik solved most of them! I could always disturb him in his office, even in the moments that he had a lot of other work to do. Marnik was always early in the lab and sometimes, the first thing I did in the morning was going to his office with a problem. His opinions were necessary to continue another day. I also want to thank him because he gave me the opportunity to work some months on the BlueGene project. Besides this professional part, I liked him very much personally. His funny jokes were welcome to continue another day. Stephanie De Vos was my neighbor at our desk for four years. As we had a the same project for a different species, we were frequently confronted with the same problems and so, we did learn a lot from each other. We first had to share the AFLP technology and then the next generation sequencing. Working together with her was really a nice experience to me. Our monthly meetings in ‘de sterre’ were very useful. We talked about our problems and progression with Wim Vyverman, Koen Sabbe, Marnik Vuylsteke, Marie Huysman, Valerie Devos and sara Moeys. These meeting were a source of new ideas and were very useful in my progression. I would like to thank Pieter Vanormelingen and Olga Chepurnova for learning me the Seminavis culturing techniques. Ilse vercauteren learned the AFLP technology to me and as she was our lab manager, we could ask her all lab related things. I also would like to thank Tine Verstraete who assisted me in the BSA-AFLP procedure. Valerie Devos learned the radio-active AFLP procedure to me. Marie huysman assisted me in a lot of things and in the phylogenetic analyses. Wilson Ardiles assisted in sequencing and a lot of people from the PSB learned little things to me, necessary in this research. After the linkage mapping, a new era of next generation sequencing analyses started for me. A lot of learning was needed in this new challenging field. Yao-Cheng Lin, Stephane Rombauts, Lieven Sterck and Frederik Delaere from the bio-informatics and IT groups teached me the first steps in Linux and in a lot of programs. The VIB bio-informatics training and service facility (BITS) organized bio-informatics courses and whenever a problem arose, Joachim Jacob from BITS was always willing to help. In the lab, as members of our little corner (Stephanie, myself, Annelies en then Sara), we helped each other a lot and of course, we also had a lot of fun together. In special I would like to thank the members of the PAE, the Quantitative Genomics and Seed Development group for all the nice moment in and outside the lab, for the nice teambuildings and all the fun we made. For people at my home front, I will switch to dutch. Mijn familie, schoonfamilie en vrienden wil ik graag bedanken. Zonder de onvoorwaadelijke steun van mijn ouders, die me alle kansen gegeven hebben, had ik hier nu niet gestaan. Ook mijn broers en schoonzussen wil ik bedanken voor alle steun. Mijn nichtjes, Laura en Julie, zijn erbij gekomen tijdens mijn doctoraat. Jullie komst was enorm mooi. Tot slot, Shanah, wil ik jou bedanken. Jij bent het mooiste geschenk van deze laatste vier jaar. Gedurende mijn doctoraat heb ik jou leren kennen en zijn we getrouwd. Onze onvoorwaardelijke liefde voor mekaar is het mooiste waar ik ooit van kon dromen. Ives Contents Abbreviations ........................................................................................................................... 11 1 ................................................................................................................................................ 15 Introduction .............................................................................................................................. 15 Diatoms: general description ................................................................................................ 15 Ecological importance ...................................................................................................... 15 Main features and evolution ............................................................................................. 16 Current and potential applications of diatoms .................................................................. 24 The diatom cell and life cycle .............................................................................................. 25 General features ................................................................................................................ 25 Variation in reproductive features .................................................................................... 26 Diatom model organisms for molecular biology .................................................................. 28 Whole genome models ..................................................................................................... 28 Genetic transformation in diatoms ................................................................................... 29 Seminavis robusta – a new model to study the diatom life cycle .................................... 29 Sex determination in eukaryotes .......................................................................................... 32 General features ................................................................................................................ 32 Evolution of sex chromosomes ........................................................................................ 33 Variation in sex differentiation in eukaryotes .................................................................. 34 Main techniques used ........................................................................................................... 39 AFLP technology ............................................................................................................. 39 Genetic linkage mapping .................................................................................................. 41 Whole genome sequencing (WGS) .................................................................................. 42 Bulked segregant analysis (BSA) ..................................................................................... 43 Aims and thesis outline ........................................................................................................ 43 Literature cited ..................................................................................................................... 44 2 ................................................................................................................................................ 53 Linkage mapping identifies the mating type determining region as a single locus in the pennate diatom Seminavis robusta ........................................................................................... 53 Abstract ................................................................................................................................ 54 Introduction .......................................................................................................................... 55 Materials and methods ......................................................................................................... 57 Production of a F1 mapping population ............................................................................ 57 Phenotyping: determining the mating type of the F1 progeny ......................................... 57 DNA extraction of S. robusta cultures ............................................................................
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