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Leaf Morpho-Anatomical Evaluation of Echeveria Cultivars using Different Fixation Methods
Tran My Khanh Thi Ha,Cabahug-Braza Raisa Aone M.,임기병,황윤정 한국원예학회 2024 원예과학기술지 Vol.42 No.2
Echeveria succulent plants are among the most diverse ornamental plants belonging to the Crassulaceae family, showing increased demand in horticultural markets owing to their rosette formation and high adaptability to water-stress environments. Owing to this level of demand and its desirable qualities, several studies have attempted to define environmental adaptation for this plant as well as its phenotypes and taxonomy, usually through leaf morpho-anatomical research. Leaf anatomical analysis has been widely employed to study these relationships; however, in the case of Echeveria, there are several barriers owing to their unique leaf structure, meaning that relatively few studies have been conducted. Several techniques use chemical fixation to investigate leaf anatomy; however, there are no universal methods due to equipment constraints and a lack of comparative studies. In this study, different fixation methods using formaldehyde, ethanol, and methanol were used to visualize the leaf morpho-anatomical structures of 15 Echeveria cultivars under light and scanning electron microscopy (LM and SEM), whereas freehand sectioning was used to obtain transverse sections. Each fixative had a different effect on each cultivar despite belonging to the same genus. Among the three fixation methods, methanol provided better preservation of the cell morphology as observed using LM and SEM. In addition, freehand sectioning is a cost-effective method that requires skill and patience, limited and affordable materials, and can be generally used without any sophisticated equipment. The results of this study provide a benchmark for future research employing simple and effective methods for related species that possess succulent leaf characteristics.
Cytogenetic Analysis and Genome Size Estimation of Korean Native Veronica Taxa
My Khanh Thi Tran Ha,Samantha S Sevilleno,Raisa Aone M Cabahug,Ji-Hun Yi,Hye Jin Oh,Hyuck Hwan Kwon,Sang-Yong Kim,Yoon-Jung Hwang 한국화훼학회 2022 화훼연구 Vol.30 No.2
Veronica L., the largest genus in the family Plantaginaceae, is widespread in various habitats. Due to their long-blooming flowers, Veronica species have high horticultural value as indoor potted, garden, and landscape plants. Furthermore, Veronica plants are extremely important owing to their notable diversity in habitat usage, ploidy level, and evolution. Several native taxa, which are of key interest in breeding programs and phylogenetic studies, have been identified in Korea. The genome sizes and chromosomal characteristics are basic cytogenetic features of all taxa, and their knowledge is a prerequisite when commencing genome sequencing projects. It can provide essential information for cytogenetic, taxonomic, phylogenetic, and evolutionary studies. Thus, cytogenetic analysis and genome size estimation of seven Veronica taxa native to Korea were conducted in this study. Fluorescence in situ hybridization (FISH) karyotype analysis and chromosome counting was conducted using metaphase chromosomes probed with 5S and 45S rDNA. Nuclear DNA content and genome size were determined using flow cytometry. FISH karyotype analysis revealed a common number of 5S loci and varying 45S signals that create distinctive rDNA distribution patterns in each taxon. The results indicated that the seven investigated Veronica taxa have calculated genome sizes (1C values) ranging from 517.1 to 862.0 Mbp. This study is the first to report the chromosome number and karyomorphology of seven Veronica taxa native to Korea, as well as the use of rDNA markers for identifying individual chromosomes. These findings contribute to the crucial understanding the genomic characteristics of species within the genus Veronica, serve as a basis for studying Veronica phylogeny and evolution, and provide valuable information for future breeding programs.
LD<SUB>50</SUB> Evaluation in Echeveria Cultivars Induced by Chemical Mutagens
Ha Tran Thi My Khanh,Raisa Aone M. Cabahug,Ki-Byung Lim,Yoon-Jung Hwang 한국원예학회 2021 한국원예학회 학술발표요지 Vol.2021 No.10
Mutation breeding has been applied by plant breeders worldwide with the expectation of improvement on the quality of ornamental plants and other crop varieties in respect to their morphological and physiological attributes such as unique visual quality, change in color, increased pests/diseases resistance. The chemical mutation is one of the effective tools in mutation breeding that is widely used as it is inexpensive with high indices of mutations induction. However, the concept of exposure chemical mutagens does to achieve ideal results is not unified, lethal dose 50% (LD50) study is accessed to determine which concentration would enable regeneration survival by 50%. Among abundant ornamental plants, Echeveria is chosen for this study due to their unique characteristics such as their color and exceptional leaf formation, and drought tolerance which has gain demand in horticulture communities. Four chemical mutagens, namely colchicine, ethyl methanesulfonate (EMS), methyl methanesulfonate (MMS), and oryzalin were used to induce three Echeveria cultivars (E. ‘Doterang’, E. ‘Elegans’, and E. ‘Hyaliana’ at different concentrations for colchicine and oryzalin (0.2%, 0.4%, 0.6%, 0.8%, 1%), and EMS and MMS (0.1%, 0.2%, 0.3%, 0.4%, 0.5%), and soaked at different times (3, 6, 9, and 12 hr). The results revealed that chemical mutagens have significantly affected the survival rates and have varied per cultivar. Leaf cuttings treated in oryzalin had the highest survival rate about 85%, followed by colchicine of about 65% at lower concentrations and at 1% concentration with 55%. The planting materials treated with EMS and MMS were significantly affected by soaking durations and concentrations. The results showed that using 0.2% concentration at short time duration at 3 hr of EMS produced 58% survival rates, however, those dipped in MMS had less than half survival count from the total treated leaf cuttings. The results of this study highly provided preliminary knowledge to mutation breeding programs for other succulent varieties or other related crops.
Phenotype and Ploidy Analysis of the Colchicineinduced M₁ Generation of Echeveria Species
Raisa Aone M. Cabahug,Ha Tran Thi My Khanh,Ki-Byung Lim,Yoon-Jung Hwang 한국원예학회 2020 원예과학기술지 Vol.38 No.4
Mutation breeding is an important tool for developing new cultivars in horticulture. Among the many methods of mutation breeding, chemical mutation is highly effective and can be performed easily. Compared to natural breeding methods, higher mutation rates and the faster induction of desirable characteristics have been reported with the use of chemical mutagens. Succulents have recently gained popularity because of their unique geometrical shapes and their ability to survive with minimal watering. Succulents that have peculiar shapes and colors demand higher prices. In this study, we used colchicine, a chemical mutagen, and tested its application on three Echeveria succulent species. A phenotypic evaluation was conducted on the mutant succulents produced from the application of colchicine on propagated leaf cuttings. Phenotypic evaluation included plant parameters and morphological analysis. Ploidy analysis was conducted to confirm the effects of the mutagen treatments. In all selected Echeveria species, the use of colchicine produced mutant species that varied significantly from those of the control; however, treatment concentration and duration varied per species. The phenotypic evaluation revealed that colchicine-mutated plants exhibited compactness, with mutants being generally taller with a thicker but shorter plant diameter compared to that of the control. Mutated plants exhibited prominent changes in color for the a<SUP>*</SUP> and b<SUP>*</SUP> values. Similarly, changes in leaf shape were observed and were evident at their apexes. These morphological changes are attributed to the change in ploidy level, which was confirmed through stomata and ploidy analysis. Larger stomata size was accompanied by lower stomata density. Based on the flow cytometry analysis, mutated succulents exhibited a 2x-4x complex.