Tileye Feyissa

Recent achievements obtained by chloroplast transformation

Chloroplasts play a great role for sustained wellbeing of life on the planet. They have the power and raw materials that can be used as sophisticated biological factories. They are rich in energy as they have lots of pigment-protein complexes capable of collecting sunlight, in sugar produced by photosynthesis and in minerals imported from the plant cell. Chloroplast genome transformation offers multiple advantages over nuclear genome which among others, include: integration of the transgene via homologus recombination that enables to eliminate gene silencing and position effect, higher level of transgene expression resulting into higher accumulations of foreign proteins, and significant reduction in environmental dispersion of the transgene due to maternal inheritance which helps to minimize the major critic of plant genetic engineering. Chloroplast genetic engineering has made fruit full progresses in the development of plants resistance to various stresses, phytoremediation of toxic metals, and production of vaccine antigens, biopharmaceuticals, biofuels, biomaterials and industrial enzymes. Although successful results have been achieved, there are still difficulties impeding full potential exploitation and expansion of chloroplast transformation technology to economical plants. These include, lack of species specific regulatory sequences, problem of selection and shoot regeneration, and massive expression of foreign genes resulting in phenotypic alterations of transplastomic plants. The aim of this review is to critically recapitulate the latest development of chloroplast transformation with special focus on the different traits of economic interest.

Somatic embryogenesis of a coffee (Coffea arabica L.) hybrid using leaf explants

Summary Coffea arabica is one of the most traded agricultural commodities in the World, and the major export commodity of Ethiopia. Improvement of this crop requires the development of hybrid varieties that possess desirable traits. The production and distribution of hybrid coffee is difficult due to the high costs required for manual crossing and maintenance. The use of somatic embryogenesis is an effective means of propagation. A protocol for somatic embryogenesis of hybrid C. arabica from leaf explants is described here. Leaf explants collected from healthy 1-year-old seedlings were used. The highest percentage of callus induction was observed from explants cultured on MS medium containing 1.0 mg l–1 2,4-dichlorophenoxyacetic acid (2,4-D) in combination with 2.0 mg l–1 6-benzylaminopurine (BAP). Embryogenic calli were obtained from leaf explants cultured on MS medium supplemented with 0.05 mg l–1 kinetin in combination with 0.1 mg l–1 indole-3-butyric acid (IBA). The highest number of embryos that germinated per explant (14.0 ± 1.7) was obtained on MS medium supplemented with 2.0 mg l–1 BAP in combination with 0.5 mg l–1 gibberellic acid (GA3). The maximum number of roots per plantlet (3.0 ± 1.0) was obtained on half-strength MS medium containing 0.5 mg l–1 indole-3-butyric acid (IBA). Acclimatisation of plantlets was achieved with a survival rate of 96.9%. This is an important achievement as acclimatising embryo-derived plantlets had been the most challenging feature of coffee tissue culture in Ethiopia. This protocol will enable us to multiply and supply large numbers of high quality hybrid coffee seedlings to farmers engaged in coffee production.

Characterization of Ethiopian Sweet Sorghum Accessions for 0Brix, Morphological and Grain Yield Traits

Abstract The improvement of sweet sorghum (Sorghum bicolor) for biofuel traits is getting more attention globally due to its sugar-rich stalk that can be used as a renewable energy product. An understanding and proper assessment of biofuel-related traits in sweet sorghum crop is an important step toward the development of superior cultivar. The study was conducted to characterize sweet sorghum collections of Ethiopia for 0Brix degree, some morphological and grain yield traits. A total of 180 accessions were collected and evaluated in two environments. The result revealed wide variability among the accessions collected from different regions. Collections from northern part of Ethiopia (Wello and Tigray) showed significantly high 0Brix mean value while collections from the rest of the regions (Hararge, West Shewa, East Wollega and Gojam) expressed lower 0Brix mean value but higher biomass for stalk sugar yield. Most of the accessions showed higher 0Brix mean value, which is similar to globally known sweet sorghum cultivars. Accessions with high 0Brix degree combined with high biomass traits like stalk diameter and plant height will be an ideal germplasm to be used for biofuel production. Hence, it is imperative to utilize these accessions in sweet sorghum breeding program in order to develop superior sugar-rich sweet sorghum cultivars.

Detection and characterization of Maize chlorotic mottle virus and Sugarcanemosaic virus associated with maize lethal necrosis disease in Ethiopia: an emerging threat to maize production in the region

Maize Lethal Necrosis Disease (MLND) develops from synergistic co-infection by Sugarcane mosaic virus and Maize chlorotic mottle virus. The disease causes symptoms ranging from leaf tissue mottling and malformed ears to premature plant death. Information on the diversity and transmission of those viruses are limited in Ethiopia. No attempts were made to characterize and identify SCMV and MCMV strains of Oromia and Benishangul-Gumuz regions. The aim of this research was to study MLND responsible viruses via serological and molecular techniques. A total of 174 symptomatic maize leaf samples were collected. Using Double Antibody Sandwich-ELISA (DAS-ELISA), 43 samples showed strong reaction for MCMV antibody and only 14 samples were clearly positive for SCMV. Reverse transcriptase polymerase chain reaction (RT-PCR) technique revealed the presence of MCMV and SCMV in most samples. Analysis of the genetic diversity of MCMV and SCMV by sequencing the coat protein of all isolates shows strong homology with previous East African isolates deposited in the GenBank. The phylogenetic analysis indicated that the virus isolates are highly similar with each other. In conclusion, this study unequivocally confirmed the association of MCMV and SCMV with MLND in the investigated areas and for the first time provided information on the genetic variability by sequence analysis of several Ethiopian samples. Since the risk posed by MLND in the investigated area is high, there is an urgent need for developing integrated management options for controlling the disease.

Genetic diversity study of common bean (Phaseolus vulgaris L.) germplasm from Ethiopia using inter simple sequence repeat (ISSR) markers

Phaseolus vulgaris L. (family Leguminosae), is a leguminous crop widely distributed in all parts of the world. In Ethiopia, common bean is cultivated as a source of protein for local consumption and for export. Mostly, it grows in the warm and lowland areas of the country. The aim of this research was to investigate the genetic diversity of P. vulgaris accessions from Ethiopia. Twelve accessions of P. vulgaris were used to estimate the genetic diversity among and within accessions using inter simple sequence repeat (ISSR) markers. Seven selected ISSR primers yielded 69 reproducible bands from 96 individuals studied. All the loci were found to be polymorphic. The total genetic diversity (H) and Shannon’s diversity information index (I) for the entire accessions showed 0.35 and 0.53, respectively. Individuals from Shinile site showed the highest level of gene diversity (H = 0.29, I = 0.43); while the lowest variability was shown by accessions from Metekel (L) (H = 0.10, I = 0.15). Variation within accessions was higher (62.55%) as compared to that of inter accessions (37.45%) based on AMOVA. In PCoA, majority individuals of Metekel (L) tended to form separate group. The result of the study confirmed the presence of genetic diversity that can be exploited to improve the productivity. This calls for a conserted efforts in the collection, conservation and sustainable use of P. vulgaris . Keywords: Genetic diversity, ISSR, Phaseolus vulgaris African Journal of Biotechnology , Vol 13(31) 3638-3649

Micropropagation of Plectranthus edulis (Vatke) Agnew from meristem cultur

Plectranthus edulis (Vatke) Agnew is a tuber-bearing food crop in Ethiopia. However, its productivity is hampered by shortage of pathogen-free planting materials. Therefore, the objective of this study was to develop micropropagation protocol for this plant using meristem to produce clean planting materials. Mersitems were collected from apical and axillary shoots from Holeta and Welayta areas and cultured on MS medium containing gibberellic acid (GA 3 ) (1.0 mg l -1 ), α-naphthalene acetic acid (NAA) (0.1 mg l -1 ) in combination with 6-benzylaminopurine (BAP) (0.1, 0.5, 1.0, 2.0 and 5.0 mg l -1 ). Shoots were multiplied on MS medium containing 0.1, 0.5, or 1.0 mg l -1 of BAP or 0.5, 1.0, 2.0 or 3.0 mg l -1 of Kinetin alone or their combination with 0.05 or 0.1 mg l -1 NAA. In vitro and ex vitro rooting was performed using different types of auxins followed by acclimatization. The highest percentage of shoots initiated from collected mersitem at Holeta (73%) was obtained on MS medium containing 1.0 mg l -1 BAP, 1.0 mg l -1 GA 3 and 0.1 mg l -1 NAA. The highest shoot number explants -1 (7.2) was obtained on MS medium containing 1.0 mg l -1 Kinetin and 0.1 mg l -1 NAA, whereas the highest root number shoot -1 (6.2) was obtained from ex vitro . All plants derived from Holeta and 96.7% of those from the Welayta survived after acclimatization. These results provided rapid and reproducible conditions for propagation of relatively pathogen-free planting material of this plant. Keywords: In vitro propagation, meristem culture, micropropagation, Plectranthus edulis (Vatke) Agnew, shoots multiplication African Journal of Biotechnology , Vol 13(31) 3682-3688

Leaf and seed yield of jute mallow (Corchorus olitorius L.) accessions under field conditions for two consecutive growing seasons

ABSTRACT Jute mallow is a nutritious leafy vegetable used by many people in Africa. The purpose of this study was to select accessions with high leaf yield and seed yield. We investigated seven agronomic traits that are related to leaf yield and seed yield and how they correlate with each other. The results indicated significant differences among the accessions in all traits. Leaf fresh weight ranged from 18.3 to 121.3 g/plant in accessions TOT 6747 and TOT 8532, respectively. The highest variability between the accessions was also observed in seed yield. It ranged from 1.0 g/plant in accession TOT 7980 to 35.5 g/plant in TOT 7866. Eight other accessions had a seed yield of 1.0 g/plant. Significant and positive correlations were observed between leaf fresh weight and leaf dry weight (r = 0.84), leaf area (r = 0.33), and number of leaves (r = 0.40). A significant correlation was observed in seed yield and weight of pods per plant (r = 0.83). The evaluation of these agronomic traits for leaf and seed yield in this germplasm has revealed useful information for breeders in their efforts to improve the yield as well as selection of accessions with good agronomic traits.

Characterization of Morphological Diversity of Jute Mallow (Corchorus spp.)

Jute mallow is a traditional leaf vegetable that is an important part of daily diet for the majority of people in rural areas in sub-Saharan Africa. Here we employed quantitative and qualitative phenotypic traits to assess the morphological diversity of 90 accessions using univariate and multivariate analyses. Field experiments were conducted for two seasons to identify accessions suitable for leaf yield. The accessions were significantly variable in all traits. Highest variability among accessions was found in harvest index, biomass yield, and weight of 1000 seeds. The traits that significantly correlated with biomass yield include plant height (), petiole length (), primary branches (), and number of leaves per plant (). Principal component analysis showed that the first five PCs with eigenvalues ≥1 explained 72.9% of the total variability in the accessions. Pods per plant, primary branches, secondary branches, and number of leaves per plant accounted for highest variability in PC1. Cluster analysis grouped the accessions into five major clusters mainly based on their origin. Thus, the collection displayed high variation in morphological traits, particularly those related to leaf yield. These accessions are therefore useful in breeding for the improvement of the crop and germplasm management.

Current trends in genetic manipulations to enhance abiotic and biotic stresses in tobacco

Environmental stresses, both abiotic and biotic, are the main factors reducing crops productivity. Biotic stresses caused by fungi, viruses, bacteria and insects are the major threats. However, abiotic stresses have more adverse effects on crop yield and crop survival. Hitherto, tolerant plants were mainly produced by classical breeding techniques. Success in breeding for better adapted varieties to abiotic and biotic stresses depends on the concerted efforts of various research domains including plant and cell physiology, molecular biology, genetics and breeding. However, such process is time consuming. The production of transgenic plants by genetic engineering allows a much faster progress. This review focuses on stress tolerance on tobacco. Tolerance to salt, drought, water stress, low and high temperature, heavy metals, pathogens and pests were enhanced via overexpression of specific stress related genes from different sources. Keywords: Overexpression, genetic en g ineerin g , abiotic factors, biotic factors African Journal of Biotechnolo g y, Vol 13(21) 2095-2102

Meristem culture of selected sweet potato (Ipomoea batatas L. Lam.) cultivars to produce virus-free planting material

Summary Sweet potato (Ipomoea batatas L. Lam.) is a staple root crop, consumed as a carbohydrate-rich food source in many tropical countries. The production and productivity of sweet potato is limited by several factors, mainly virus infections. This study was conducted to produce virus-free sweet potato planting material using meristem culture. Meristems without leaf primordia were excised from four virus-positive field-grown local sweet potato cultivars ‘Bellela’, ‘Temesgen’, ‘LO-323’, and ‘Zapallo’. Meristems were cultured in Murashige and Skoog (MS) medium supplemented with different combinations and concentrations of 6-benzylaminopurine (BAP), α-naphthaleneacetic acid (NAA), and gibberellic acid (GA3) for shoot initiation. ‘Bellela’ and ‘Temesgen’ exhibited 100% shoot initiation on 5 mg l–1 BAP, 0.01 mg l–1 NAA, and 1 mg l–1 GA3, although the shoots were bushy. ‘LO-323’ showed 90% shoot initiation with the best shoot quality on 2 mg l–1 BAP, 0.01 mg l–1 NAA, and 1 mg l–1 GA3. Shoot tips and nodes from in vitro-grown plantlets were cultured on BAP (0, 0.5, 1.0, or 2.0 mg l–1) or on BAP (0, 0.5, 1.0, or 2.0 mg l–1) + kinetin (Kin; 0, 0.1, 0.5 or 1.0 mg l–1) for multiple shoot formation. The best shoot proliferation was obtained on MS + 0.5 mg l–1 BAP + 0.5 mg l–1 Kin. In vitro-grown shoots were cultured on MS medium containing 0.0, 0.01, 0.1, 0.5, or 1.0 mgl–1 indole-3-butyric acid (IBA). The best root formation was obtained on IBA-free medium. The plantlets thus produced were tested for the presence of ten sweet potato viruses [Sweet potato chlorotic stunt virus (SPCSV), Sweet potato feathery mottle virus (SPFMV), Sweet potato mild mottle virus (SPMMV), Sweet potato chlorotic fleck virus (SPCFV), Sweet potato caulimo-like virus (SPCaLV), Sweet potato mild speckling virus (SPMSV), C-6 (a flexuous rod virus), Sweet potato latent virus (SwPLV), Sweet potato virus G (SPVG), and Cucumber mosaic virus (CMV)] using nitrocellulose membrane enzyme-linked immunosorbent assay (NCM-ELISA). Ninety-nine percent of plantlets were free from all viruses.