Genetic analysis of the arable weeds Arabidopsis thaliana and Viola arvensis from agricultural fields with different use-histories [Elektronische Ressource] / Robert O

Genetic analysis of the arable weeds Arabidopsis thaliana and Viola arvensis from agricultural fields with different use-histories [Elektronische Ressource] / Robert O'Neill

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Genetic analysis of the arable weeds Arabidopsis thaliana and Viola arvensis from agricultural fields with different use-histories Robert O’Neill Contents Contents List of Figures vi List of Tables vii List of Terms and Abbreviations x Introduction 1 2 Literature Review 2 2.1 Agricultural Intensification 2 2.2 Biodiversity 3 2.3 Arable Weeds and Their Role in Biodiversity 4 2.4 Genetic Diversity and Biodiversity 5 2.5 Population Gentics 6 2.6 Detcting Gentic Diversity 8 2.6.1 Measures of Genetic Diversity 9 2.7 Microsatellites (Simple Sequence Repeats – SSRs) 10 2.7.1 Microsatellite Models of Evolution 12 2.7.2 Population Genetics Statistics for Microsatellites 13 2.8 Amplified Fragment Length Polymorphism 16 3 Materials and Methods 19 3.1 Sampling 19 3.1.1 Arabidopsis thaliana (L.) HEYNH. 19 3.1.2 Viola arvensis (MURR.) 2 3.1.3 Galeopsis tetrahit 23 3.2 Sampling Locations 24 3.2.1 Sampling Technique 25 3.2.2 Plant Material for Genetic Analysis 26 3.3 Molecular Marker Techniques 27 iiContents 3.3.1 DNA Extraction 27 3.3.2 Quantification 28 3.3.3 Microsatellite Analysis 3.2.4 AFLP Analysis 30 3.2.5 Gel Electrophoresis 33 3.

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Genetic analysis of the arable
weeds Arabidopsis thaliana and
Viola arvensis from agricultural
fields with different use-histories



Robert O’Neill
Contents
Contents

List of Figures vi

List of Tables vii

List of Terms and Abbreviations x

Introduction 1

2 Literature Review 2
2.1 Agricultural Intensification 2
2.2 Biodiversity 3
2.3 Arable Weeds and Their Role in Biodiversity 4
2.4 Genetic Diversity and Biodiversity 5
2.5 Population Gentics 6
2.6 Detcting Gentic Diversity 8
2.6.1 Measures of Genetic Diversity 9
2.7 Microsatellites (Simple Sequence Repeats – SSRs) 10
2.7.1 Microsatellite Models of Evolution 12
2.7.2 Population Genetics Statistics for Microsatellites 13
2.8 Amplified Fragment Length Polymorphism 16

3 Materials and Methods 19
3.1 Sampling 19
3.1.1 Arabidopsis thaliana (L.) HEYNH. 19
3.1.2 Viola arvensis (MURR.) 2
3.1.3 Galeopsis tetrahit 23
3.2 Sampling Locations 24
3.2.1 Sampling Technique 25
3.2.2 Plant Material for Genetic Analysis 26
3.3 Molecular Marker Techniques 27
iiContents
3.3.1 DNA Extraction 27
3.3.2 Quantification 28
3.3.3 Microsatellite Analysis
3.2.4 AFLP Analysis 30
3.2.5 Gel Electrophoresis 33
3. Staistcal Analysi 34
Genetic Variability 34
Hardy-Weinberg Expectations 35
Linkage Disequilibrium 35
Analysis of Molecular Variance 35
Bottleneck Detection 35
Binary Data Analysis 36
Principal Coordinate Analysis and Clustering 36
Rates of Migration 37
Mantel Test 37

4 Results 38
4.1 Sampling 38
4.2 Arabidopsi thalina 41
4.2.1 Molecular Markers 41
Microsatellite Markers and Allele Frequencies 41
AFLP Markers 47
4.2.2 Within Population Genetic Diversity 47
4.2.3 Population Differentiation and Genetic Distance 54
AMOVA analysis 54
Population Genetic Distances 5
Bottleneck Detection 59
Linkage Disequilibrium 61
4.2.4 Principle Coordinate Analysis (PCA) 61
4.2.5 Cluster Analysis 65
4.2.6 Mantel Test 6
4.2.7 Migration Rates 68
iiiContents
4.2.8 Estimation of θ 71

4.3 Viola rvensi 73
4.3.1 Viola arvensis markers 73
4.3.2 Within-Population Genetic Diversity 73
4.3.3 Population Differentiation and Genetic Distance 75
AMOVA analysis 75
Population Genetic Distances 75
4.3.4 Principle Coordinate Analysis (PCA) 77
4.3.5 Cluster Analysis 79
4.3.6 Mantel Test 79

5 Discussion 81
5.1 Sampling 81
5.2 Abundance 82
5.3 rabidopsi thalina 83
Within-population genetic diversity 83
Hardy-Weinberg Equilibrium (HWE) 86
Among-Population Genetic Relationships 87
Principle Coordinate and Cluster Analysis 90
Mantel Test 90
Bottleneck Detection 91
Ecology 92

5.4 Viola arvensis 94
Within-Population Genetic Diversity 94
Among-Population Genetic Diversity
Principle Coordinate and Cluster Analysis 95
Mantel Test 95
Ecology 96

ivContents
6 Conclusions and Recommendations 97

7 Summary 9

8 Zusammenfassung 105

9 Bibliography 112

Appendices 125
Appendix I 126
Appendix II 131 III 136

Acknowledgements 139

vList of Figures
List of Figures

Fig. 1: Arabidopsis thaliana (L.) Heynh. 19
Fig. 2: Geographical distribution of A. thaliana (L.) Heynh. 21
Fig. 3: Viola arvensis 22
Fig. 4a-4g: Allele frequency distributions for A. thaliana locus:
a) nga59 43
b) nga63 43
c) nga111 44
d) nga128 4
e) nga139 45
f) nga151
g) nga168 46
Fig. 5a-5e: Relationship between A. thaliana sample size and:
a) H 51 exp
b) H 51 obs
c) NA 51
d) NA 51 e
e) NA /NA 51 e
Fig. 6a-6c: Relationship between A. thaliana sample size and:
a) gen diversity 53
b) F 53 ST
c) R 53 ST
Fig. 7: Relationship between M and sample size for A. thaliana 61
Fig. 8a-8b: Principle coordinate analysis of A. thaliana using microsatel-
lites data by:
a) regime 63
b) location 63
Fig. 9a-9b: Principle coordinate analysis of A. thaliana using
AFLP data by:
a) regime 64
b) location 64
viList of Figures
Fig. 10a-10c: Mantel test for all A. thaliana subpopulations using:
a) F values from microsatellite data 53 ST
b) R 53 ST
c) AFLP dat 53
Fig. 11: Relationship between sample size and gene diversity
for V. arvensis 74
Fig. 12a-12b: Principle coordinate analysis of V. arvensis using
AFLP data by:
a) regime 78
b) location 78
Fig. 13: Mantel test for all V. arvensis subpopulations 80
Fig. 14: Cluster analysis of A. thaliana individuals using AFLP data 126
Fig. 15: Cluster analysis of individuals using
microsatellites data 131
Fig. 16: Cluster analysis of A. thaliana136
viiList of Tables
List of Tables

Tab. 1: Average distance between sampling sites 24
Tab. 2:Sampling sites 25
Tab. 3: Reagents used for DNA extraction 28
Tab. 4: Microsatellite primers used for A. thaliana analysis 29
Tab. 5: Thermocycler programme for PCR amplification of
microsatellites 29
Tab. 6: AFLP primer sequences for +1-amplification on
A. thaliana and V. arvensis 30
Tab. 7: AFLP Protocol for +1-amplification 31
Tab. 8: Thermocycler programme for +1-amplification 31
Tab. 9: AFLP +3-primer combinations used for V. arvensis 32
Tab. 10: AFLP +2-primer combinations used for A. thaliana 32
Tab. 11: AFLP Protocol for +3-amplification 32
Tab. 12: Thermocycler programme for +3-amplification 33
Tab. 13: Solutions used for PAGE 34
Tab. 14: Number of mature plants extracted from soil samples 38
Tab. 15: Number of A. thaliana plants analysed by location 39
Tab. 16: Number of V. arvensis 40
Tab. 17: Number of A. thaliana and V. arvensis plants analysed
by regime 40
Tab. 18: Microsatellite loci statistics for A. thaliana 42
Tab. 19: AFLP primer statistics for A. thaliana 47
Tab. 20: Population statistics by regime for A. thaliana 50
Tab. 21: Population statistics by subpopulation for A. thaliana 50
Tab. 22: Gene diversity averaged over all loci for
populations by regime 52
Tab. 23: Gene diversity averor A. thaliana
subpopulation s 53


viiiList of Tables
Tab. 24: AMOVA results for A. thaliana using:
a) pairwise difference (microsatellite data) 55
b) sum of squared size difference (microsatellite data) 55
c) pairwise difference (AFLP data) 55
Tab. 25: Genetic distance by regime for A. thaliana using
microsatellite data 56
Tab. 26: Genetic distance by regime for A. thaliana using
AFLP data 56
Tab. 27: Genetic distance between subpopulations of A. thaliana
using microsatellite data 57
Tab. 28: Genetic distance between subpopulations of A. thaliana
using AFLP data 57
Tab. 29: Bottleneck detection for A. thaliana populations 60
Tab. 30: Migration rate estimates from microsatellite data
for A. thaliana populations by regime 70
Tab. 31: Migration rate estima
for A. thaliana subpoulations 70
Tab. 32: θ values for A. thaliana populations by regime 72 Hom
Tab. 33: θ values for subpopulations 72 Hom
Tab. 34: AFLP primer statistics for V. arvensis 73
Tab. 35: Gene diversity averaged over all loci for V. arvensis
populations by regime 74
Tab. 36: Gene diversity averor V. arvensis
subpopulation s 74
Tab. 37: AMOVA results for V. arvensis 75
Tab. 38: Genetic distance by regime for V. arvensis 76
Tab. 39: Genetic distance between subpopulations of V. arvensis 76
Tab. 40: Colour code used for sample identification in dendrograms 125



ixList of Terms and Abbreviations
List of Terms and Abbreviations

IBD – Identical-By-Descent

IIS – Identical-In-State

Homoplasy - alleles that are identical-in-state, not identical-by-descent

SMM – Stepwise Mutation Model

IAM – Infinite Allele Model

F = H – H / H ST t s t
where H represents the expected level of heterozygosity in a subpopula-s
tion, and H the expected level of heterozygosity if all subpopulations were t
pooled together (i.e. in the total population).

d-F - The average number of different alleles between individuals within ST
a population

R = S – S / S ST w
where S and S are the average sum of squares of the difference in allele w
size within a subpopulation and for the entire population, respectively

d-R - the average squared difference in allele size (measured as mi-ST
crosatellite product length) between individuals within a population

Singletons – a unique AFLP band (occurring only once throughout all sam-
ples)

M - the ratio of number of alleles (k) to range in allele size (r) for any
given population
x