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(Bromeliaceae) in Den Südamerikanischen Anden

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(Bromeliaceae) in Den Südamerikanischen Anden Phylogeographie und Populationsgenetik der Arten der Gattung Fosterella (Bromeliaceae) in den südamerikanischen Anden Dissertation Natascha Wagner Phylogeographie und Populationsgenetik der Arten der Gattung Fosterella (Bromeliaceae) in den südamerikanischen Anden Dissertation zur Erlangung des akademischen Grades eines Doktors der Naturwissenschaften (Dr. rer. nat.) angefertigt in der Abteilung Morphologie und Systematik der Pflanzen, Institut für Biologie, Fachbereich 10/ Mathematik und Naturwissenschaften, Universität Kassel von Natascha Wagner Kassel, 2012 Als Dissertation vom Fachbereich 10 der Universität Kassel angenommen am: 13. Dezember 2012 Betreuer: Prof. Dr. Kurt Weising Promotionskommission: Prof. Dr. Kurt Weising (1. Gutachter) Prof. Dr. Pierre Ibisch (Beisitzer) Prof. Dr. Georg Zizka (2. Gutachter) Prof. Dr. Rüdiger Wagner (Beisitzer) Datum der Disputation: 26. Februar 2013 Inhaltsverzeichnis Inhaltsverzeichnis 1 Einleitung .............................................................................................. 1 1.1 Eine kurze Übersicht über die Erdgeschichte Südamerikas..................................... 1 1.2 Bromeliaceae ............................................................................................................ 3 1.3 Fosterella................................................................................................................. 5 1.3.1 penduliflora-Gruppe ........................................................................................ 10 1.3.2 micrantha-Gruppe............................................................................................ 14 1.4 Ziele dieser Arbeit ................................................................................................... 17 2 Material und Methoden ....................................................................... 19 2.1 Material.................................................................................................................... 19 2.1.1 Pflanzenmaterial für vergleichende Sequenzierung ........................................ 19 2.1.2 Pflanzenmaterial für populationsgenetische Untersuchungen (AFLPs).......... 19 2.2 Methoden................................................................................................................. 26 2.2.1 DNA-Isolation ................................................................................................. 26 2.2.1.1 Verwendete Reagenzien, Materialien und Geräte ................................... 26 2.2.1.2 Durchführung........................................................................................... 28 2.2.2 Polymerase-Kettenreaktion (PCR) ................................................................. 29 2.2.3 Agarosegelelektrophorese................................................................................ 30 2.2.4 Aufreinigung der PCR-Produkte ..................................................................... 31 2.2.5 Sequenzierung.................................................................................................. 31 2.2.5.1 Verwendete Reagenzien, Materialien und Geräte.................................... 31 2.2.5.2 Durchführung........................................................................................... 32 2.3 Test und Auswahl geeigneter DNA-Abschnitte für vergleichende Sequenzierung . 33 2.3.1 Plastidäre Marker............................................................................................. 33 2.3.1.1 Test potenzieller Kandidaten ........................................................................ 33 2.3.1.2 Charakterisierung der ausgewählten Chloroplastenloci rpl32-trnL und rps16-trnK.............................................................................................. 34 2.3.1.3 Amplifikation der plastidären Marker .......................................................... 35 2.3.2 Nukleäre single-copy Loci............................................................................... 36 2.3.2.1 Test potenzieller Kandidaten ................................................................... 36 2.3.2.2 Auswahl der verwendeten nukleären Loci................................................... 38 2.3.2.3 Amplifikation der nukleären Loci ................................................................ 39 I Inhaltsverzeichnis 2.4 Phylogenetische Auswertung und Analysen............................................................ 41 2.4.1 Indelkodierung ................................................................................................. 41 2.4.2 Neighbour Joining-Analyse ............................................................................. 41 2.4.3 Haplotypennetzwerke....................................................................................... 41 2.4.4 Maximum Parsimony-Analyse (MP) ............................................................... 42 2.4.5 Maximum Likelihood-Analyse (ML) .............................................................. 42 2.4.6 Bayes´sche Analyse ......................................................................................... 43 2.4.7 Datierte Phylogenien........................................................................................ 44 2.4.8 Biogeographische Analysen............................................................................. 45 2.5 AFLPs – Amplified Fragment Length Polymorphisms........................................... 47 2.5.1 Prinzip der Methode......................................................................................... 47 2.5.2 Verwendete Reagenzien und Materialien ........................................................ 48 2.5.3 Restriktion und Ligation .................................................................................. 48 2.5.4 Präselektive Amplifikation............................................................................... 49 2.5.5 Selektive Amplifikation ................................................................................... 50 2.5.6 Wiederholung misslungener Reaktionen ......................................................... 51 2.5.7 Manuelle Editierung der AFLP-Rohdaten und Normalisierung ...................... 51 2.6 Populationsgenetische Analysen der AFLP-Daten.................................................. 52 2.6.1 Neighbour Joining-Analyse ............................................................................. 52 2.6.2 NeighbourNet-Analyse .................................................................................... 52 2.6.3 Hauptkoordinaten-Analyse (PCoA)................................................................. 52 2.6.4 Manteltest......................................................................................................... 53 2.6.5 AMOVA........................................................................................................... 53 2.6.6 Structure-Analyse............................................................................................. 53 3 Ergebnisse............................................................................................ 55 3.1 Plastidäre Marker..................................................................................................... 55 3.1.1 Test und Auswahl der Chloroplastenmarker.................................................... 55 3.1.2 Phylogenetische Analysen ............................................................................... 57 3.1.2.1 Analysen der plastidären Loci.................................................................. 57 3.1.2.2 Topologie der Phylogenien....................................................................... 58 3.1.3 Datierte Phylogenie.......................................................................................... 64 3.1.4 Biogeographische Analysen............................................................................. 65 3.1.5 Haplotypennetzwerke....................................................................................... 67 II Inhaltsverzeichnis 3.1.5.1 penduliflora-Gruppe ................................................................................ 67 3.1.5.2 micrantha-Gruppe.................................................................................... 69 3.2 Nukleäre Marker...................................................................................................... 71 3.2.1 Test und Auswahl der nukleären low-/single-copy Marker ............................ 71 3.2.2 Phylogenetische Analysen............................................................................... 76 3.2.2.1 Phylogenien basierend auf NIA-i3........................................................... 76 3.2.2.2 Phylogenien basierend auf PHYC............................................................ 81 3.3 AFLPs - Amplified Fragment Length Polymorphisms ........................................... 84 3.3.1 penduliflora-Gruppe ........................................................................................ 84 3.3.1.1 AMOVA................................................................................................... 85 3.3.1.2 Neighbour Joining-Analyse ..................................................................... 88 3.3.1.3 NeighbourNet-Analyse ............................................................................ 89 3.3.1.4 Structure-Analyse..................................................................................... 90 3.3.1.5 PCo-Analyse und Manteltest ..................................................................
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