The Transcriptomic and Genomic Architecture of Acrididae Grasshoppers Dissertation To Fulfil the Requirements for the Degree of “Doctor of Philosophy” (PhD) Submitted to the Council of the Faculty of Biological Sciences of the Friedrich Schiller University Jena by Bachelor of Science, Master of Science, Abhijeet Shah born on 7th November 1984, Hyderabad, India 1 Academic reviewers: 1. Prof. Holger Schielzeth, Friedrich Schiller University Jena 2. Prof. Manja Marz, Friedrich Schiller University Jena 3. Prof. Rolf Beutel, Friedrich Schiller University Jena 4. Prof. Frieder Mayer, Museum für Naturkunde Leibniz-Institut für Evolutions- und Biodiversitätsforschung, Berlin 5. Prof. Steve Hoffmann, Leibniz Institute on Aging – Fritz Lipmann Institute, Jena 6. Prof. Aletta Bonn, Friedrich Schiller University Jena Date of oral defense: 24.02.2020 2 Table of Contents Abstract ........................................................................................................................... 5 Zusammenfassung............................................................................................................ 7 Introduction ..................................................................................................................... 9 Genetic polymorphism ............................................................................................................. 9 Lewontin’s paradox ....................................................................................................................................... 9 The evolution of genome size ................................................................................................. 11 Computational tools for de novo TE and repetitive DNA discovery ............................................................ 15 De novo discovery, classification and annotation of transposable elements ............................................. 16 The Acrididae Grasshoppers ................................................................................................... 17 Gene expression analysis and de novo transcriptome assembly .............................................. 22 Orthopteran transcriptomes ....................................................................................................................... 22 Transcript analysis using RNA-seq............................................................................................................... 23 De novo transcriptome assembly ................................................................................................................ 24 Overview of the Oases assembler ............................................................................................................... 25 Overview of Trinity assembler .................................................................................................................... 26 Overview of SOAPdenvo-trans transcriptome assembler ........................................................................... 27 Challenges and overview ........................................................................................................ 28 Objectives and Structure ........................................................................................................ 30 Manuscript Overview ..................................................................................................... 31 Manuscript I ................................................................................................................................................ 31 Manuscript II ............................................................................................................................................... 31 Manuscript III .............................................................................................................................................. 32 Manuscript I ................................................................................................................... 33 Manuscript II .................................................................................................................. 44 Manuscript III ................................................................................................................. 55 Abstract ................................................................................................................................. 56 Introduction .......................................................................................................................... 57 Results ................................................................................................................................... 60 Repetitive content and genome size ........................................................................................................... 60 Characterization of repeat content within species ..................................................................................... 61 Divergences within clusters of transposable elements ............................................................................... 62 Variation in repeat content across species ................................................................................................. 62 Intra-specific differences in Gomphocerus sibiricus .................................................................................... 63 Species differences explored by cluster painting ........................................................................................ 64 Discussion .............................................................................................................................. 64 Methods ................................................................................................................................ 68 Species and sample collection .................................................................................................................... 68 Genome size determination by flow cytometry ......................................................................................... 68 High throughput sequencing and short read pre-processing ..................................................................... 69 Phylogenetic analysis .................................................................................................................................. 70 De novo repeat identification ..................................................................................................................... 70 Iterative repeat identification and filtering................................................................................................. 71 Repeat content estimation ......................................................................................................................... 72 Joint repeat clustering and comparison across species .............................................................................. 72 3 Cluster annotation ...................................................................................................................................... 73 Ancestral state reconstruction .................................................................................................................... 74 Comparative analysis of the migratory locust............................................................................................. 74 Data access .................................................................................................................................................. 75 Tables and Figures ................................................................................................................. 75 Figure legends ............................................................................................................................................. 75 Figure 1 ....................................................................................................................................................... 77 Figure 2 ....................................................................................................................................................... 78 Figure 3 ....................................................................................................................................................... 79 Figure 4 ....................................................................................................................................................... 80 Figure 5 ....................................................................................................................................................... 81 Figure 6 ....................................................................................................................................................... 82 Supplementary Material for Manuscript III ............................................................................. 83 Figure S1 ...................................................................................................................................................... 83 Figure S2 ...................................................................................................................................................... 84 Figure S3 ...................................................................................................................................................... 85 Figure S4 .....................................................................................................................................................