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The Identification of Functional, Sequestered, Symbiotic Chloroplasts

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The Identification of Functional, Sequestered, Symbiotic Chloroplasts University of South Florida Scholar Commons Graduate Theses and Dissertations Graduate School 2006 The identification of functional, sequestered, symbiotic chloroplasts in Elysia clarki: A crucial step in the study of horizontally transferred, nuclear algal genes Nicholas E. Curtis University of South Florida Follow this and additional works at: http://scholarcommons.usf.edu/etd Part of the American Studies Commons Scholar Commons Citation Curtis, Nicholas E., "The identification of functional, sequestered, symbiotic chloroplasts in Elysia clarki: A crucial step in the study of horizontally transferred, nuclear algal genes" (2006). Graduate Theses and Dissertations. http://scholarcommons.usf.edu/etd/2496 This Dissertation is brought to you for free and open access by the Graduate School at Scholar Commons. It has been accepted for inclusion in Graduate Theses and Dissertations by an authorized administrator of Scholar Commons. For more information, please contact [email protected]. The Identification of Functional, Sequestered, Symbiotic Chloroplasts in Elysia clarki: A Crucial Step in the Study of Horizontally Transferred, Nuclear Algal Genes by Nicholas E. Curtis A thesis submitted in partial fulfillment of the requirements for the degree of Doctor of Philosophy Department of Biology College of Arts and Sciences University of South Florida Major Professor: Sidney K. Pierce, Jr., Ph.D. Clinton J. Dawes, Ph.D. Kathleen M. Scott, Ph.D. Brian T. Livingston, Ph.D. Date of Approval: June 15, 2006 Keywords: Bryopsidales, kleptoplasty, sacoglossan, rbcL, chloroplast symbiosis Penicillus, Halimeda, Bryopsis, Derbesia © Copyright 2006, Nicholas E. Curtis Note to Reader The original of this document contains color that is necessary for understanding the data. The original dissertation is on file with the USF library in Tampa, Florida. Table of Contents List of Tables...................................................................................................................v List of Figures ............................................................................................................... vi Abstract ......................................................................................................................... ix Preface........................................................................................................................... xi Introduction.....................................................................................................................1 Chapter 1: A Morphological and Molecular Comparison Between Elysia crispata and a New Species of Kleptoplastic Sacoglossan Sea Slug (Gastropoda: Opisthobranchia) from the Florida Keys, USA......................................5 Introduction..........................................................................................................5 Materials and Methods .........................................................................................7 Animals and Habitats................................................................................7 Florida Keys .................................................................................7 Virgin Islands................................................................................8 Radular Preparation ..................................................................................9 Chloroplast Localization and Morphology ................................................9 Chloroplast Identifications......................................................................10 PCR and Sequencing ..............................................................................11 Comparison of Mitochondrial Gene Sequences.......................................12 i Results ...............................................................................................................14 Taxonomy ..............................................................................................14 Etymology ..................................................................................14 Paratypes.....................................................................................14 Distribution.................................................................................14 Comparison Between E. clarki n. sp. and E. crispata..............................16 External Morphology ..................................................................17 Dorsal Surface Venation .............................................................22 Chloroplast Location and Morphology ........................................26 Chloroplast Identification............................................................31 Discussion..........................................................................................................34 Chapter 2: Molecular Phylogeny of the Bryopsidales (Ulvophyceae, Chlorophyta) Based on a Partial Sequence of the Chloroplast Encoded, Large Subunit Ribulose Bisphosphate Carboxylase/Oxygenase Gene.......................40 Introduction........................................................................................................40 Materials and Methods .......................................................................................43 Specimen Sampling and Outgroup Selection...........................................43 DNA Extraction......................................................................................44 PCR and Sequencing ..............................................................................45 Phylogenetic Analysis.............................................................................46 Results ...............................................................................................................47 Exhaustive Maximum Parsimony............................................................48 Bayesian Analysis ..................................................................................49 ii Discussion..........................................................................................................51 Chapter 3: The Symbiotic Chloroplasts in the Sacoglossans Elysia clarki are from Several Algal Species ................................................................................57 Introduction........................................................................................................57 Materials and Methods .......................................................................................59 Animals and Algae .................................................................................59 DNA Extractions ....................................................................................60 PCR and Sequencing ..............................................................................60 Electron Microscopy...............................................................................61 Feeding...................................................................................................62 Results ...............................................................................................................63 rbcL Sequence Identity...........................................................................63 Electron Microscopy...............................................................................63 Discussion..........................................................................................................71 Chapter 4: Newly Metamorphosed Elysia clarki Juveniles Feed on and Sequester Chloroplasts from Algal Species Different from Those Utilized by Adult Slugs............................................................................................75 Introduction........................................................................................................75 Materials and Methods .......................................................................................77 Sea Slugs and Algae ...............................................................................77 Culture of Elysia clarki...........................................................................80 Juvenile Feeding.....................................................................................81 Electron Microscopy...............................................................................81 iii DNA Extractions ....................................................................................83 PCR and Sequencing ..............................................................................84 Results ...............................................................................................................86 Juvenile Feeding Preferences..................................................................86 Ultrastructure of Symbiotic Chloroplasts ................................................87 Molecular Identification of Elysia clarki Chloroplasts ............................94 Discussion..........................................................................................................96 Conclusions .................................................................................................................100 References ...................................................................................................................103 About the Author................................................................................................End Page iv List of Tables Table 1 Comparison of the lengths of radular teeth from Virgin Islands slugs, wild Keys slugs and
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