Population Genetic Analysis of Neolithic to Bronze Age Human Remains from Trentino-Alto Adige (Northern Italy)
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POPULATION GENETIC ANALYSIS OF NEOLITHIC TO BRONZE AGE HUMAN REMAINS FROM TRENTINO-ALTO ADIGE (NORTHERN ITALY) DISSERTATION ZUR ERLANGUNG DES AKADEMISCHEN GRADES DOKTOR DER NATURWISSENSCHAFTEN (DR. RER. NAT.) AM FACHBEREICH BIOLOGIE DER JOHANNES GUTENBERG-UNIVERSITÄT MAINZ VORGELEGT VON ANGELA GRAEFEN GEB. AM 13.12.1975 IN CHERTSEY/GROßBRITANNIEN MAINZ, 2019 Tag der mündlichen Prüfung: 20. April 2020 Table of Contents i TABLE OF CONTENTS 1 INTRODUCTION AND BACKGROUND ...................................................... 3 1.1 The Neolithisation of Europe ................................................................................ 3 1.2 Bronze Age Europe and the Pontic steppe migration ............................................. 7 1.3 Relative male/female contribution during Neolithic/Bronze Age migrations ......... 9 1.4 Prehistory of South Tyrol and the Adige Valley .................................................. 12 1.4.1 Mesolithic ............................................................................................... 12 1.4.2 Neolithic ................................................................................................. 14 1.4.3 Copper Age (Chalcolithic)....................................................................... 16 1.4.4 Bronze Age ............................................................................................. 20 1.5 Population genetics of South Tyrol and the Adige Valley.................................... 23 1.6 Aim and objective ............................................................................................... 26 2 MATERIAL AND METHODS ................................................................... 28 2.1 Sample material .................................................................................................. 28 2.1.1 Characteristics of sampled sites ............................................................... 31 2.1.2 Prehistoric samples.................................................................................. 32 2.1.3 Chronological overview .......................................................................... 47 2.2 Genetic markers .................................................................................................. 50 2.2.1 Mitochondrial DNA ................................................................................ 50 2.2.2 Functional and phenotypic markers ......................................................... 51 2.2.3 Y-chromosomal DNA ............................................................................. 54 2.3 Ancient DNA analyses ........................................................................................ 59 2.3.1 General ancient DNA anticontamination and authentication measures ..... 59 2.3.2 Selection and preparation of bone and tooth samples ............................... 61 2.3.3 DNA extraction ....................................................................................... 62 2.3.4 PCR primer design .................................................................................. 63 2.3.5 Polymerase chain reaction (PCR) ............................................................ 65 2.3.6 Multiplex PCR ........................................................................................ 66 2.3.7 Gel electrophoresis .................................................................................. 67 2.3.8 DNA purification (PCR product) ............................................................. 67 2.3.9 Cycle sequencing .................................................................................... 68 2.3.10 Purification II (cycle sequencing product) ............................................... 68 2.3.11 Capillary electrophoresis ......................................................................... 69 2.3.12 Sequence analysis ................................................................................... 69 2.3.13 Scoring system ........................................................................................ 70 2.3.14 Reference data sets .................................................................................. 71 2.3.15 Statistical analyses (mtDNA) .................................................................. 85 3 RESULTS ............................................................................................... 91 3.1 Amplification and sequencing success ................................................................ 91 3.2 Authentication & reproducibility ........................................................................ 93 3.2.1 Discrepancies due to sampling errors....................................................... 93 3.2.2 Scoring system results ........................................................................... 100 3.3 Contamination rate ........................................................................................... 103 3.4 Sequence data ................................................................................................... 104 3.4.1 Mitochondrial DNA sequences .............................................................. 105 3.4.2 Nuclear DNA sequences........................................................................ 112 3.4.3 Y-chromosomal SNPs ........................................................................... 117 3.5 Population genetic analyses .............................................................................. 119 3.5.1 Group structuring .................................................................................. 119 3.5.2 Statistical analyses ................................................................................ 125 Table of Contents ii 4 DISCUSSION ........................................................................................ 146 4.1 Population history of ancient Trentino populations ........................................... 146 4.1.1 Neolithic ............................................................................................... 146 4.1.2 Copper Age ........................................................................................... 147 4.1.3 Bronze Age ........................................................................................... 153 4.2 Phenotypic loci ................................................................................................. 159 4.2.1 Lactase persistence (-13.910*T) ............................................................ 159 4.2.2 Eye colour ............................................................................................. 161 4.3 Routes of Neolithisation ................................................................................... 162 4.4 Autosomal link between Tyrolean Iceman and Sardinia .................................... 164 4.5 Site continuity in the Trentino region from the Mesolithic to the Bronze Age ... 166 4.6 Links with modern populations ......................................................................... 168 4.6.1 Haplotype sharing versus FST values ...................................................... 175 4.7 Links between Ancient Trentino and the Tyrolean Iceman ................................ 176 5 CONCLUSIONS .................................................................................... 178 5.1 Zusammenfassung ............................................................................................ 185 5.2 Riassunto .......................................................................................................... 187 6 APPENDIX ........................................................................................... 189 6.1 Instruments and reagents ................................................................................... 189 6.2 Abbreviations ................................................................................................... 192 6.3 List of figures ................................................................................................... 193 6.4 List of tables ..................................................................................................... 194 6.5 Supplementary information on CD.................................................................... 195 6.5.1 Supplement S1: List of samples with sequencing results........................ 195 6.5.2 Supplement S2: Reference sample sets (ancient) ................................... 195 6.5.3 Supplement S3: Reference sample sets (modern) ................................... 195 6.5.4 Supplement S4: Haplotype frequencies (ancient) ................................... 196 6.5.5 Supplement S5: Haplotype frequencies (modern) .................................. 196 6.5.6 Supplement S6: Full scoring results ....................................................... 196 6.5.7 Supplement S7: FST values .................................................................... 196 6.5.8 Supplement S8: Positive controls yielding sequence .............................. 197 6.5.9 Supplement S9: Samples from lab personnel ......................................... 197 6.5.10 Alignment reports ................................................................................. 197 6.6 References ........................................................................................................ 198 Introduction and Background 3 1 INTRODUCTION AND BACKGROUND 1.1 The Neolithisation of Europe The spread of the Neolithic to central Europe, leading to a shift in human subsistence from hunting and gathering to an agricultural