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GENETIC DIVERSITY and ECO-GEOGRAPHICAL DISTRIBUTION of Eleusine SPECIES COLLECTED from ETHIOPIA

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GENETIC DIVERSITY and ECO-GEOGRAPHICAL DISTRIBUTION of Eleusine SPECIES COLLECTED from ETHIOPIA African Crop Science Journal, Vol. 22, No. 1, pp. 45 - 58 ISSN 1021-9730/2014 $4.00 Printed in Uganda. All rights reserved © 2014, African Crop Science Society GENETIC DIVERSITY AND ECO-GEOGRAPHICAL DISTRIBUTION OF Eleusine SPECIES COLLECTED FROM ETHIOPIA L. DAGNACHEW, SANTIE DE VILLIERS1, T. SEWALEM, M. DIDA2, F. MASRESHA3, W. KIMANI1 and T. KASSAHUN Department of Microbial, Cellular & Molecular Biology, Addis Ababa University, Ethiopia 1International Crop Research Institute for Semi-Arid Tropics (ICRISAT), Nairobi, Kenya 2Department of Botany and Horticulture, Maseno University, Nairobi, Kenya 3Department of Plant Biology and Biodiversity Management, Addis Ababa University, Ethiopia Corresponding author: [email protected] (Received 8 November, 2013; accepted 5 February, 2014) ABSTRACT Eleusine is a small grass genus with three basic chromosome numbers (x=8, 9 and 10) and comprises of eight species including E. coracana subsp. coracana, (finger millet), which is an important subsistence crop in Africa and India. Research on these species could assist the development of high yielding and multiple stress tolerant variety(s) of the cultivable species, and also guide development of strategic genetic resource management and utilisation of the genus. A total of 72 accessions, sampled from five major species, E. coracana (including both E. coracana subsp. coracana and E. coracana subsp. africana), E. intermedia, E. indica, E. multiflora and E. floccifolia were analysed for genetic variation and inter-relationships using 20 microsatellite markers. All the SSR markers displayed high genetic polymorphism, with polymorphic information content ranging from 0.46 (UGEP110) to 0.91 (UGEP66). A total of 286 alleles were observed with an average of 14.3 alleles per locus. Classic F-statistics revealed the highest intra-specific polymorphism recorded for E. africana (32.45%), followed by E. coracana (16.83%); implying that genetic polymorphism is higher in the cultivable subspecies and its wild relatives, than the other species. Allelic frequency based inter-species genetic distance analysis, showed wider genetic distance between E. indica and E. multiflora (0.719); a narrow genetic distance between E. coracana sub- species africana and E. coracana subspecies coracana (0.3297). The weighted neighbor joining-based clustering revealed that the majority of the accessions in a species share strong similarity and are grouped together than do accessions of inter species. Key Words: Eleusine, microsatellite, polymorphism RÉSUMÉ Eleusine est une herbe avec trois nombres de chromosomes de base (x=8, 9 et 10) et comprend huit espèces dont E. coracana subsp. coracana, (finger millet), qui est une culture de subsistence importante en Afrique et en Inde. La recherche sur ces espèces pourrait aider dans le développement des variétés d’espèces cultivables à rendement élevé et de tolérance aux stress multiples, et guider le développement de la gestion des ressources génétiques stratégiques et l’utilisation du genus. Un total de 72 accessions échantillonnées de cinq espèces majeurs à savoir E. coracana (incluant E. coracana subsp. coracana et E. coracana subsp. africana), E. intermedia, E. indica, E. multiflora et E. floccifolia étaient analysées pour variation génétique et relations mutuelles utilisant 20 marqueurs microsatellites. Tous les marquers SSR ont manifesté un polymorphisme génétique élevé, avec un contenu d’information polymorphique allant de 0.46 (UGEP110) à 0.91 (UGEP66). Un total de 286 allèles était observé avec une moyenne de 14.3 allèles par locus. Les statistiqies classiques F ont révélé le polymorphisme intraspécifique le plus élevé enregistré pour le E. africana (32.45%), suivi de E. coracana (16.83%), ce qui implique que le polymorphisme génétique est le plus élevé dans les sous espèces cultivables et ses homologues sauvages que les 46 L. DAGNACHEW et al. autres espèces. L’analyse de la fréquence allélique de la distance génétique entre espèces a montré une plus large distance génétique entre E. indica et E. multiflora (0.719); une étroite distance génétique entre les sous espèces Africana de E. coracana et les sous espèces coracana de E. coracana (0.3297). La pondération des groupements a révélé que la majorité des accessions au sein d’une espèce partage une forte similarité et sont groupées ensemble en comparaison aux accessions des intra-espèces. Mots Clés: Eleusine, microsatellite, polymorphisme INTRODUCTION Neves et al., 2005) and gene flow between subspecies (Dida et al., 2008). This is mainly The genus Eleusine Gaertn. comprises of eight caused by co-occurrence of the two E. coracana species, among which the cultivable E. coracana subspecies in the same crop fields where they subsp. coracana (finger millet) is the most are cultivated and as weeds, for instance in important subsistence crop of Africa and India. Western and North Western Ethiopia (Tsehay, On the other hand, E. indica (goose grass) is 2012). Several authors also confirmed that the categorised as one of the most problematic weeds cultivated subspp. coracana (finger millet) was in the world (Holm et al., 1977; Phillips, 1995; domesticated through natural and artificial Devarumath et al., 2005). The genus is selection, from the wild type finger millet, subspp. characterised by three basic chromosome africana (Hilu and De Wet, 1976; Hilu and numbers of x=8, 9 and 10; whereby E. intermdia, Johnson, 1992; Dida et al., 2008). E. indica, E. floccifolia and E. tristachya are Bisht and Mukai (2002) indicated that E. diploids with 2n=2x=18; E. multiflora is a diploid indica, E. tristachya, E. floccifolia and E. with 2n= 2x=16; E. jaegeri is also diploid with intermedia, E. coracana, subspecies coracana 2x=2n=20 (Devarumath et al., 2005). The and E. coracana subspecies africana are closely tetraploid, Eleusine spp., comprise of E. related and that there is free genetic flow between coracana subsp coracana and subsp. africana, them. Eleusine multiflora was reported as with 2n=4x=36, and E. kigeziensis with 2n=38, significantly different morphologically and which all probably have allopolyploid origins genetically from other species (Neves, 2011). The involving two diploid species with x=9 and 10 distribution, population, genetic variation and (Bisht and Mukai, 2002; Neves, 2011). preferential adaptation of species can be Dida and Devos et al. (2006) categorised the influenced by altitude gradients that comprise of species of the genus into a A genome group, an assemblage of environmental factors such as, comprising of E. indica and E. tristachya and a climatic and other edaphic factors (Korner, 2007; B genome group comprising three species, Ohsawa and Ide, 2008). Research on those species namely E. floccifolia, E. intermedia and E. might bring a novel gene that can be introgrssed multiflora. Since the evolution of the tetraploid into the cultivable E. coracana subsp. coracana species involved both A and B genome groups; to develop good yielder and/or stress tolerant E. coracana subspecies coracana (finger millet), varieties, particularly against the devastating E. coracana subspecies africana and E. finger millet blast disease. Furthermore, studying kigeziensis belong to both genome groups (Dida the eco-geographical distribution of plant species and Devos, 2006). All the species are of African is essential for efficient genetic resource origin, except E. tristachya, which is native to collection, appropriate in-situ germplasm South America (Devarumath et al., 2005; Neves conservation/management and elucidating the et al., 2005). taxonomy, evolution and origin of the species The frequent and naturally occurring (Bekele, 1985; Demissie and Bjornstrand, 1996). hybridisation between cultivated E. coracana Therefore, this study was aimed at characterising subsp. coracana and its wild relative E. coracana the genetic diversity, inter- and intra- species subsp. africana gave rise to many morphological relationships and the eco-geographical patterning intermediates (De Wet et al., 1984; Phillips, 1995; of the different species collected from various Genetic diversity and eco-geographical distribution of Eleusine species 47 regions of Ethiopia as a basis for germplasms published SSR markers (Table 3) for finger millet collection and conservation. (Dida et al., 2007). All forward primers contained an M13-tag (5’- CACGACGTTGTAAAACGAC - MATERIALS AND METHODS 3’) on the 5’ end that was fluorescently labeled to allow detection of amplification products Plant materials. A total of 72 accessions of five (Shuelke, 2000). PCR amplification was performed different species of the genus Eleusine (E. in 10 µl in 384 well microtitre plates. Each reaction coracana (including E. coracana subsp. comprised of 1 x PCR buffer (20 mM Tris-HCl, pH coracana and E. coracana subsp. africana), 7.6; 100 mM KCl; and 0.1 mM EDTA. Other E.intermedia, E.indica, E. multiflora and E. components included 1 mM DTT; 0.5% (w/v) floccifolia) were included in the study (Table 1). Triton X-100; 50% (v/v) glycerol), 2 mM MgCl2, Collections were done in part of Addis Ababa 0.16 mM dNTPs, 0.16 µM fluorescent labeled administrative region, Amhara regional state, M13-forward primer, 0.04 µM forward primer, 0.2 Benishangul Gumuz regional state, Oromia µM reverse primer, 0.2 units of Taq DNA regional state, Southern Nations Nationalities and polymerase (Sib Enzyme Ltd, Russia) and 30 ng Peoples Regional State of Ethiopia (Table 1). Some of template DNA. PCR reactions were performed additional accessions were originally collected
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