Title Page
Contents
LIST OF ABBREVIATIONS 17
GENERAL ABSTRACT 18
GENERAL INTRODUCTION 21
LITERATURE REVIEW 24
A. Background of Succulents 24
ⅰ Characterization of Succulent Plants 24
ⅱ. Cultivation and Importance of Ornamental Succulents 25
ⅲ. Controversy in the phylogenetic relationship of Echeveria species 26
B. Leaf anatomical analysis 27
ⅰ. Fundamental tissues 28
ⅱ. Stomatal Characteristics 30
ⅲ. Vascular tissue arrangement 32
C. Cytogenetic information on the Echeveria genus 34
ⅰ. Chromosome variation 34
ⅱ. Nuclear DNA content 36
D. Mutation Breeding 38
REFERENCES 40
CHAPTER 1. Leaf Morpho-Anatomical Evaluation of Echeveria Using Different Fixation Methods Observed Under Light and Scanning Electron Microscope: The Implication of Its Anatomical Structure to Their Environment 48
ABSTRACT 49
Ⅰ. INTRODUCTION 50
Ⅱ. MATERIALS AND METHODS 53
A. Plant materials 53
B. Morphological evaluation 54
C. Free-hand sectioning 54
D. Preparation for Fixation methods 54
E. Leaf micro-morphology observation 55
F. Moisture content measurement 55
G. Statistical and Spearman's correlation analysis 56
Ⅲ. RESULTS 57
A. Evaluation of fixation methods 57
B. Leaf micro-morphology 67
C. Leaf anatomical structure response to environmental conditions 74
Ⅳ. DISCUSSION 76
A. Evaluation of fixation methods 76
B. Anatomical structure and its response to environmental conditions 78
Ⅴ. CONCLUSION 82
REFERENCES 83
CHAPTER 2. Somatic Chromosome Assessment of Echeveria Succulents 87
ABSTRACT 88
Ⅰ. INTRODUCTION 89
Ⅱ. MATERIALS AND METHODS 92
A. Plant materials 92
B. Mitotic chromosome preparation 93
Ⅲ. RESULTS 93
Ⅳ. DISCUSSION 97
Ⅴ. CONCLUSION 101
REFERENCES 102
CHAPTER 3. LD₅₀ Analysis, Phenotypic Evaluation, and Cytogenetics Analysis of Colchicine-Induced Echeveria Cultivars 104
ABSTRACT 105
Ⅰ. INTRODUCTION 106
Ⅱ. MATERIALS AND METHODS 108
A. Plant materials 108
B. Experimental design and treatments 108
C. Preparation and application of chemical mutagens 108
D. Data measurement and collection 108
E. Stomata characteristics 109
F. Chromosome analysis 109
G. Nuclear DNA extraction 110
Ⅲ. RESULTS 111
A. E. 'Brave' 111
B. E. 'Peerless' 113
C. Stomata analysis 116
D. Chromosome numbers counting and flow cytometry analysis 117
Ⅳ. DISCUSSION 120
Ⅴ. CONCLUSION 123
REFERENCES 124
CHAPTER 4. Evaluation of Genome Size and Endoreduplication in Echeveria 128
ABSTRACT 129
Ⅰ. INTRODUCTION 130
Ⅱ. MATERIALS AND METHODS 133
A. Plant materials 133
B. Estimation of Nuclear DNA Content 133
C. Endoreduplication Determination 134
Ⅲ. RESULTS 135
Ⅵ. DISCUSSION 138
Ⅳ. CONCLUSION 141
REFERENCES 142
국문초록 145
CHAPTER 1 15
Table 1. Echeveria species and cultivars information. 54
Table 2. Morphological characteristics of Echeveria species and cultivars. 66
Table 3. Leaf micro-morphology observed under LM of Echeveria species and cultivars. 68
Table 4. Leaf-micromorphology parameters of Echeveria species and cultivars. 70
Table 5. Stomata characteristics of Echeveria species and cultivars. 74
Table 6. Spearman's correlation of leaf morphology, characteristics, and moisture content of Echeveria species and cultivars. 75
CHAPTER 2 15
Table 7. Echeveria species and cultivars information. 93
Table 8. Somatic chromosome numbers of Echeveria species and cultivars. 96
CHAPTER 3 15
Table 9. Survival and mutant rate (%) of Echeveria cultivars induced with colchicine (n=50). 111
Table 10. Plant parameters of mutated E. 'Brave' treated with colchicine with different concentrations and dipping times at 12 MAT. 113
Table 11. Color reading and equivalent color group based on the Royal Horticultural Society (RHS) color scheme for mutated E. 'Brave'. 113
Table 12. Plant and leaf growth parameters of E. 'Peerless' treated with colchicine. 115
Table 13. CIELAB color values of colchicine-induced mutant E. 'Peerless'. 116
Table 14. Stomata density and size of mutant Echeveria cultivars treated with colchicine. 117
Table 15. Chromosome count of colchicine induced on E. cultivars. 118
CHAPTER 4 16
Table 16. Analysis of DNA content, coefficient of variation (CV), level of endocycles, and number of cell cycles of Echeveria species and cultivars. 135
LITERATURE REVIEW 10
Fig. 1. Leaf anatomical section of E. 'Peerless' showing a single epidermis (EP) layer and followed below it is the hypodermis (HP) (100×, Scale Bar=50μm). 30
Fig. 2. Stomatal evaluation of E. 'Loy' showing: A. the anisocytic stomatal type comprising of three subsidiary cells (800×, Scale bar=10 µm); and the abundance of stomatal density as pointed... 32
Fig. 3. Leaf cross-section of Echeveria 'Chubby Bunny' showing: A. the 3D arrangement of the central vascular bundle (CVB) with dispersed peripheral vascular bundles (PVB) (0.8×, Scale... 33
CHAPTER 1 10
Fig. 4. Fifteen (15) Echeveria species and cultivars subjected for leaf anatomical evaluation: A. E. colorata E. Walther; B. E. 'Benbadis'; C. E. 'Brave'; D. E. 'Cubic Frost'; E. E. 'Dark Ice';... 53
Fig. 5.1. Leaf anatomical cross-sections fixed using A. FAA; B. Ethanol; and C. Methanol showing the epidermis and hypodermis layer of Echeveria species and cultivars: 1. E. colorata E.... 58
Fig. 5.2. Leaf anatomical cross-sections fixed using A. FAA; B. Ethanol; and C. Methanol showing the epidermis and hypodermis layer of Echeveria species and cultivars: 1. E. 'Doterang'; 2. E.... 59
Fig. 5.3. Leaf anatomical cross-sections fixed using A. FAA; B. Ethanol; and C. Methanol showing the epidermis and hypodermis layer of Echeveria species and cultivars: 1. E. 'Peerless'; 2. E.... 60
Fig. 6.1. Leaf micro-morphology of Echeveria species and cultivars observed under SEM: 1. E. colorata E. Walther; 2. E. 'Benbadis'; 3. E. 'Brave'; 4. E. 'Cubic Frost'; 5. E. 'Dark Ice' using various... 62
Fig. 6.2. Leaf micro-morphology of Echeveria species and cultivars observed under SEM: 1. E. 'Doterang'; 2. E. 'Fire Pillar'; 3. E. 'Glam Pink'; 4. E. 'Loy'; 5. E. 'Milk Rose' using various fixation methods: A.... 63
Fig. 6.3. Leaf micro-morphology of Echeveria species and cultivars observed under SEM: 1. E. 'Peerless'; 2. E. 'Silhouette'; 3. E. 'Snow Bunny'; 4. E. 'Tippy'; and 5. E. 'Viyant' using various fixation... 64
Fig. 7.1. Leaf micro-morphology of Echeveria species and cultivars observed under LM: A. E. colorata E. Walther; B. E. 'Benbadis'; C. E. 'Brave' ; D. E. 'Cubic Frost'; E. E. 'Dark... 67
Fig. 7.2. Leaf micro-morphology of Echeveria species and cultivars observed under LM: J. E. 'Milk Rose'; K. E. 'Peerless'; L. E. 'Silhouette'; M. E. 'Snow Bunny'; N. E. 'Tippy'; and O. E.... 68
Fig. 7.3. Leaf micro-morphology of Echeveria species and cultivars observed under SEM: 1. E. 'Peerless'; 2. E. 'Silhouette'; 3. E. 'Snow Bunny'; 4. E. 'Tippy'; and 5. E. 'Viyant' using...[내용없음] 11
Fig. 8.1. Transverse section of Echeveria species and cultivars observed under LM: A. E. colorata E. Walther; B. E. 'Benbadis'; C. E. 'Brave'; D. E. 'Cubic Frost'; E. E.'Dark Ice'; F. E. 'Doterang';... 71
Fig. 8.2. Transverse section of Echeveria species and cultivars observed under LM: J. E. 'Milk Rose'; K. E. 'Peerless'; L. E. 'Silhouette'; M. E. 'Snow Bunny'; N. E. 'Tippy'; and O. E. 'Viyant'... 72
CHAPTER 2 12
Fig. 9. Twenty-eight Echeveria species and cultivars, 1. E. colorata E. Walther; 2. E. hyaliana E. Walther; 3. E. lilacina Kimnach & Moran; 4. E. setosa; 5. E. 'Benbadis'; 6. E. 'Brave'; 7. E.... 92
Fig. 10. Somatic chromosome number counting of Echeveria species and cultivars, 1. E. colorata E. Walther 2n=54, 2. E. hyaliana E.Walther 2n=ca. 104, 3. E. lilacina Kimnach & Moran 2n=54, 4. E. setosa 2n=54, 5. E. 'Benbadis' 2n=ca. 89, 6. E. 'Brave' 2n=76, 7. E. 'Clui' 2n=50, 8. E. 'Cubic Frost' 2n=ca. 106, 9. E. 'Doterang' 2n=53, 10. E. 'Fire Lip' 2n=... 95
CHAPTER 3 13
Fig. 11. Sample comparison of the A. control and B. mutant plants exhibiting morphological changes in the: 1. Whole plant; 2. Leaf apex shape of E. 'Brave' . 112
Fig. 12. Sample comparison of the A. control and B. mutant plants exhibiting morphological changes in the: 1. whole plant; 2. leaf apex shape of E. 'Peerless'. 115
Fig. 13. Mitotic chromosome numbers counting for colchicine-induced E. 'Brave' mutants: A. control (2n=〉 75); B. 0.4% + 3 h (2n=〉 206) (400× magnification, Scale bar=10 µm). 118
Fig. 14. Mitotic chromosome numbers counting for colchicine-induced E. 'Peerless' A. control (2n=〉 110); B. 0.2% + 3 h (2n=〉 206); and C. 0.4% + 3h (2n=〉 215) (400× magnification,... 118
Fig. 15. Estimation of the nuclear DNA content of E. 'Brave': A. control and B. 0.4% + 3 h, showing corresponding peaks to nuclei 2C and 4C with an external standard test using Glycine max... 119
Fig. 16. Estimation of the nuclear DNA content of E. 'Peerless': A. control, B. 0.2% + 3 h, and C. 0.4% + 3 h showing corresponding peaks to nuclei 2C and 4C with an external standard test... 119
CHAPTER 4 14
Fig. 17. Flow cytometry results showing endoreduplication cycles of the reference: A. S. burrito and succulent species and cultivars: B. E. colorata E. Walther, C. E. 'Benbadis', D. E. 'Brave', E. E. 'Cubic Frost', F. E. 'Dark Ice', G. E. 'Doterang', H. E. 'Fire Pillar', I. E. 'Glam Pink', J. E. 'Loy',... 137