Diversification of beaked Redfish, Sebastes mentella, Along a Depth Gradient in the central North Atlantic Peter Shum This thesis is presented to the School of Environment & Life Sciences, University of Salford, in fulfilment of the requirements for the degree of Ph.D. Supervised by Prof. Stefano Mariani March 2016 Beaked redfish Sebastes mentella (Travin 1951) Table of contents Table of Contents Table of Contents ................................................................................................................................ iii List of figures ........................................................................................................................................ vi List of tables ....................................................................................................................................... viii Acknowledgements ............................................................................................................................... ix Abstract ......................................................................................................................................... xii Chapter I: Introduction 1.1 Introduction ...................................................................................................................... 1 1.2 Divergence along the population-species continuum....................................................... 2 1.3 Environmental gradients .................................................................................................. 4 1.4 Visual perception ............................................................................................................. 7 1.5 Evolutionary origins ....................................................................................................... 10 1.6 Sebastes mentella ........................................................................................................... 11 1.7 Genetic structure by depth ............................................................................................. 12 1.8 Fisheries management .................................................................................................... 13 Chapter II: Three-Dimensional Post-Glacial Expansion and Diversification of an Exploited Oceanic Fish 2.1 Abstract .......................................................................................................................... 16 2.2 Introduction .................................................................................................................... 17 2.3 Materials and methods ................................................................................................... 19 2.3.1 Sample collection ................................................................................................... 19 2.3.2 Generation of molecular data ................................................................................. 22 2.3.3 Data analysis........................................................................................................... 23 2.4 Results ............................................................................................................................ 26 2.4.1 Mitochondrial DNA lineages ................................................................................. 26 2.4.2 Comparative nuclear and mitochondrial data ......................................................... 29 2.5 Discussion ...................................................................................................................... 34 2.5.1 Population structure and connectivity .................................................................... 35 2.5.2 Historical reconstruction ........................................................................................ 38 2.6 Conclusions .................................................................................................................... 40 Chapter III: The Visual Pigment in Sebastes mentella and its Significance for Habitat Depth 3.1 Abstract .......................................................................................................................... 41 3.2 Introduction ................................................................................................................... 41 iii Table of contents 3.3 Materials and Methods ................................................................................................... 43 3.3.1 Sample collection ................................................................................................... 43 3.3.2 Generation of rhodopsin sequences and analysis ................................................... 44 3.3.3 Differential bleaching and analysis ........................................................................ 45 3.4 Results ............................................................................................................................ 46 3.4.1 Rhodopsin ............................................................................................................... 46 3.4.2 Visual spectra ......................................................................................................... 47 3.5 Discussion ...................................................................................................................... 49 3.5.1 Convergent evolution to depth ............................................................................... 49 3.5.2 Spectral sensitivity ................................................................................................. 50 3.6 Conclusions .................................................................................................................... 53 Chapter IV: RNA-seq Reveals Adaptive Processes to Different Depth Environments in Beaked Redfish, Sebastes mentella 4.1 Abstract .......................................................................................................................... 54 4.2 Introduction .................................................................................................................... 54 4.3 Materials & methods ...................................................................................................... 56 4.3.1 Animals and sample collection ............................................................................... 56 4.3.2 RNA isolation, library preparation, and sequencing .............................................. 57 4.3.3 Transcriptome assembly and functional annotation ............................................... 58 4.3.4 Analysis of gene expression ................................................................................... 59 4.4 Results ............................................................................................................................ 60 4.4.1 Transcriptome assembly and annotation ................................................................ 60 4.4.2 Differential expression ........................................................................................... 63 4.4.3 Visual genes ........................................................................................................... 63 4.5 Discussion ...................................................................................................................... 66 4.5.1 Rhodopsin sensitivity ............................................................................................. 66 4.5.2 Opsin expression .................................................................................................... 67 4.5.3 Other important findings: Immune response .......................................................... 69 4.6 Conclusions .................................................................................................................... 70 Chapter V: MtDNA Approach Resolves Ambiguity of “Redfish” Sold in Europe 5.1 Abstract .......................................................................................................................... 72 5.2 Introduction .................................................................................................................... 72 5.3 Materials and methods ................................................................................................... 74 iv Table of contents 5.3.1 Sample collection ................................................................................................... 74 5.3.2 Molecular analysis .................................................................................................. 75 5.3.3 Data analysis........................................................................................................... 75 5.4 Results ............................................................................................................................ 76 5.4.1 Marker diversity ..................................................................................................... 76 5.4.2 Sequence identification .......................................................................................... 76 5.4.3 Phylogenetic analysis ............................................................................................. 79 5.5 Discussion ...................................................................................................................... 81 5.5.1 Complicating issues for DNA barcoding for recently radiating species ................ 82 5.5.2 Phylogenetic representation ..................................................................................