In another study, Propidium iodide based flow cytometry was conducted for evaluating cytogenetic variations within the Beta group for which 21 Portugal-situated populations were studied. The study revealed variation in endopolyploidy levels for B. macrocarpa and B. vulgaris subsp. maritime (Castro et al., 2013). Karyomorpho-logical study of C. endivia and C. intybus L. was done in order to reveal differences between them. rDNA probes-based FISH followed with fluorochrome staining on five accessions for each species exhibited conserved positions and rDNA-based polymorphisms in endivia and untybus species respectively (Bernardes et al., 2013). Such techniques have paved the way to solving karyotyping and taxonomical description problems of many species. However, correct modeling needs incorporation of environmental variables, molecular mechanisms and suitable karyotypic analysis for spe-ciation and genome studies. The ploidy level and genetic makeup of different root and tuber crops are given in Table 2.2.
2.2.3 Ecological Study
Features of plant ecology can prove extremely useful in its identification. A plant needs to adapt to a specific soil, water and other environmental conditions so as to facilitate its growth and proliferance. The association of the plant species with the specific surroundings and ecological niche becomes strong and distinctive, in a way restricting and limiting them.
Plants of Xanthosoma spp. in the Kulonprogo District, Yogyakarta were characterized and identified using morphological and isozyme patterns, which were further analyzed through dendogram and statistical tools like goodness of fit (Nurmiyati and Sajidan 2009). With respect to plants at different locations, Glutamate oxaloacetic transaminase (GOT) and Peroxidase (POD) isozyme banding patterns exhibited variability in contrast to other isozymes like Esterase (EST). The features adapted by the plant in response became their mark of identification and aid in taxonomical study (Givnish, 2010). The focus of this aspect covers the study of the lifespan of plants:
• Ephemerals ― short life cycle, characteristic to dry and disturbed areas;
• Annuals ― live for one growing season, lack woody stem or root/storage organs development;
• Biennials ― proliferate in two growing seasons, possess storage organs against unfavorable periods between two seasons;
• Perennials ― many seasons, develop woody parts and specialized perennating organs (i.e. bulbs and rhizomes) and their habit forms (trees, shrubs and bushes, herbs, climbers) (Wondafrash, 2008).
Study of such features narrows down the efforts needed for the identification and description of an unknown plant and increases the pace for taxonomical study. Studies on M. esculenta plants collected from the Ngawi District at three different altitudes exhibited influential effects of height on root, stem and leaf morphology (Tribadi et al., 2010). Identification of minor changes in genetic composition between taxa of a particular habitat can also be done with the help of isozymes, which prove an inexpensive method in such cases (Crawford, 2014). Nguyen et al. (1998) developed phylogenetic relationships for 84 Southeast Asia taro species through esterase isozyme polymorphism and UPGMA cluster analysis. Genetic variability as exhibited by banding patterns was found in different accessions collected from different places, such as Nepal, Vietnam, Yunnan, etc., implying a significant role of Yunnan in evolution of the species. Similarly, Kang and Chung (2000) worked on 30 Korean Hemerocallis species, genetic population structure, variability and divergence pattern with 12 isozyme loci. Higher variations with less differentiation among populations were observed in these species. Scattered distribution of these plants in focus land areas determined limited seed dispersal. The H. hakuunensis population was mainly localized to open areas, granitic or humus soils, pine-oak forest margins localized on the hillsides of the central, southern and northwestern side of the Korean Peninsula, whereas pine-oak forests harboring sandy soils situated on the Taean Gun coast (Central-western Korean Peninsula) mainly is inhabited by H. taeanensis.
2.2.4 Chemotaxonomy
Plants possess an incredible gamut of metabolites, which can be utilized as taxonomic markers and this lead to the emergence of chemotaxonomy in the 1960s. Analysis of essential oils or other targeted metabolites, assessment of chemical data among plants and use of computational data analysis for determining similarity have been a part of this approach. Flavonoids proved to be one of the successful chemotaxonomic markers for the Leguminosae family, but characteristic leaf flavonoids are yet to be developed (Rewald et al., 2012).
Tavares et al. (2014) used fruit essential oils’ chemical characterization to derive distinctness among four D. carota subspecies namely, subsp. maximus, subsp. carota, subsp. halophilus and subsp. gummifer and demonstrated that instead of considering D. carota subsp. maximus as a subspecies of D. carota, it must be placed separately as a species itself, as it singly exhibited high asarone levels in oils. Subsp. carota and subsp. gummifer contain a high geranyl acetate content, whereas the elemicin content is high in subsp. Halophilus, exhibiting possible distinguishing characters for taxonomical classification. Similarly, root specific marker molecules are prime requirements for root taxa determination by this approach but is in its infancy. Neutral cumarins or anthraquinones have also aided in root taxa and biomass determination in a few genera. New advancements have included whole metabolites study formed or present in a plant in accordance with environmental, genetic and developmental changes. Tundis et al. (2014) enlisted different essential oils present in Stachys and reviewed their role in chemosystematic studies. Iridoids, Chrysoeriol sheleton, Isoscutellarein groups and Apigenin-p-coumaroyl derivatives have been reported as Stachys chemotaxonomic markers, whereas tricetin methyl esters based glycosides act as markers for Betonica taxa, which needs to be further studied and used for other suitable applications.
2.2.5 Molecular Identification
Phenotypic methods for plant identification prove inadequate to reason out evolutionary and taxonomic relations in closely related species and thus generate a need for incorporation of molecular tools for taxonomical study. Power of resolving genetic differences, data type generated and the applicability to different taxonomic levels influence the selection of molecular techniques. DNA barcoding, Random amplified polymorphic DNA (RAPD), Amplified fragment length polymorphism (AFLP), and Microsatellites and Single nucleotide polymorphisms (SNPs) are some of the techniques employed for studying diversity and phylogenetic relationships (Arif et al., 2010).