Browsing by Author "Kiggundu, Andrew"

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    Building a bi-directional promoter binary vector from the intergenic region of Arabidopsis thaliana cab1 and cab2 divergent genes useful for plant transformation
    (African Journal of Biotechnology, 2013) Tindamanyire, Jimmy M.; Townsley, Brad; Kiggundu, Andrew; Tushemereirwe, Wilberforce; Sinha, Neelima
    The ability to express genes in a controlled and limited domain is essential to succeed in targeted genetic modification. Having tools by which to rapidly and conveniently generate constructs which can be assayed in a diverse array of plant species expedites research and end-product development. Targeting specifically green plant tissues offers an opportunity to effect changes to diverse processes such as water use efficiency, photosynthesis, predation and nutrition. To facilitate the generation of transgenes to be expressed in this domain, we created a series of plasmids called p2CABA based on the Arabidopsis thaliana chlorophyll a/b gene promoter, a single natural bidirectional promoter that can drive and express two different genes at the same time. Studies we carried out showed reporter gene, GUS expressed in leaves and stems but not in the roots, as expected since this endogenous promoter controls the expression of two photosynthetic genes in A. thaliana. We, therefore, utilized the intergenic region between the A. thaliana cab1 and cab2 divergent genes to design and construct a bidirectional promoter vector containing two multiple cloning sites and a gateway recombination cassette. This in turn will help minimize gene silencing and achieve desirable expression pattern of transgenes, a critical issue in plant genetic engineering and in this report we show their use in Medicago and tomato.
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    Corm damage caused by banana weevils Cosmopolites sordidus (Germar) collected from different banana growing regions in Uganda
    (Agriculture & Food Security, 2018) Twesigye, Charles K.; Ssekatawa, Kenneth; Kiggundu, Andrew; Tushemereirwe, Wilberforce; Matovu, Enock; Karamura, Eldad
    In this study, both healthy tissue culture plantlets and maiden suckers of the Nakitembe cultivar were used to assess the damage level variation caused by banana weevils collected from different banana growing regions. Seventy-nine (79) tissue culture plantlets and fifty (50) suckers were established in buckets in a randomized complete block design for 5 months. Ten adult weevils (5 females and 5 males) were introduced at the base of each plant, and the buckets were covered with a weevil proof mesh. Weevil damage was estimated as a percentage at 60 days after the weevil introduction by estimating the peripheral damage (PD), total cross section corm damage (XT) and above the collar damage (ACD). Results: The results showed high differences in the PD, XI, XO and XT caused by weevils from the different zones. PD and XT ranged from 4.8–50.4 to 4.2–43.8%, respectively, caused by weevils collected from Kabale and Rakai, Kabale and Wakiso, respectively, while XI and XO varied from 0.0–42.9 to 8.3–40.4%, respectively, caused by banana weevils collected from Kabale and Rakai, Kabale and Rakai, respectively. Banana weevils from Rakai caused the highest ACD of 40.4% and no such damage was caused by banana weevils collected from western Uganda. Average ACD in suckers was 19.6% and significantly higher than that in tissue culture plants (8.5%). Conclusions and recommendations: Corm damage assessment suggests the existence of banana weevil biotypes but it is recommended that follow-up studies be carried out to confirm this phenomenon.
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    Establishment of a transformation protocol for Uganda’s yellow passion fruit using the GUS gene
    (African Journal of Biotechnology, 2019) Tuhaise, Samuel; Nakavuma, Jesca L.; Adriko, John; Ssekatawa, Kenneth; Kiggundu, Andrew
    Horticulture is one of the fastest growing sectors in Uganda, exporting products worth US$100 million annually. Passion fruit (Passiflora edulis) growing and export is one of the critical contributors to this sector employing over a million farmers. However, a number of biotic and abiotic constraints have initiated widespread enterprise abandonment by farmers. Passiflora improvement efforts by conventional breeding has had limited success calling for research into alternative approaches such as genetic engineering. The study aimed at optimizing existing protocols to develop an efficient and reproducible Agrobacterium mediated transformation system to suit Uganda’s Passiflora cultivars. Agrobacterium tumefaciens strain AGL1 (OD600 of 0.5) harbouring pCAMBIA2301 containing the GUS (uidA) reporter gene was used to infect pre-cultured leaf discs. Leaf discs were then vacuum infiltrated for 1.5 min at 750 mmHg followed by a three day co-cultivation period on MS + acetosyringone (100 μml-1). Putatively transgenic yellow passion fruit shoots were induced on Murashige and Skoog (MS) selection media supplemented with benzylaminopurine (BAP) 8.9 μM, kanamycin (100 mgL-1mgl) and cefotaxime (500 mgL-1). Developed shoots were then transferred to elongation media (MS + 0.44 μM BAP) and later rooted on 5.37 μM naphthaleneacetic acid (NAA). Genetic transformation was monitored using GUS staining. A single independently transformed plant was confirmed by polymerase chain reaction (PCR), translating in a transformation efficiency of 0.456%. A viable in vitro transformation protocol for Uganda’s yellow passion fruit directly from leaf discs was developed using GUS reporter gene. Further investigations are required to improve the reported protocols transformation efficiency.
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    Experiences in sub-Saharan Africa with GM crop risk communication
    (GM crops & food, 2013) Racovita, Monica; Obonyo, Dennis Ndolo; Anguzu, Robert; Bamwenda, Gratian; Kiggundu, Andrew; Maganga, Harrison
    In tackling agricultural challenges, policy-makers in sub-Saharan Africa (SSA) have increasingly considered genetically modified (GM) crops as a potential tool to increase productivity and to improve product quality. Yet, as elsewhere in the world, the adoption of GM crops in SSA has been marked by controversy, encompassing not only the potential risks to animal and human health, and to the environment, but also other concerns such as ethical issues, public participation in decision-making, socio-economic factors and intellectual property rights. With these non-scientific factors complicating an already controversial situation, disseminating credible information to the public as well as facilitating stakeholder input into decision-making is essential. In SSA, there are various and innovative risk communication approaches and strategies being developed, yet a comprehensive analysis of such data is missing. This gap is addressed by giving an overview of current strategies, identifying similarities and differences between various country and institutional approaches and promoting a way forward, building on a recent workshop with risk communicators working in SSA.
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    Genetic diversity of aflatoxin-producing Aspergillus flavus isolated from groundnuts in selected agroecological zones of Uganda
    (BMC microbiology, 2020) Acur, Amos; Arias, Renée S.; Odongo, Steven; Tuhaise, Samuel; Ssekandi, Joseph; Muhanguzi, Dennis; Adriko, John; Buah, Stephen; Kiggundu, Andrew
    Background Aspergillus is the main fungal genus causing pre- and post-harvest contamination of groundnuts. Aspergillus flavus belongs to section Flavi, a group consisting of both the aflatoxigenic species (A. flavus, A. parasiticus and A. nomius) and non-aflatoxigenic species (A. oryzae, A. sojae and A. tamarii). Aflatoxins are food-borne toxic secondary metabolites produced by Aspergillus species, causing hepatic carcinoma and stunting in children and are the most toxic carcinogenic mycotoxins ever identified. Despite the well-known public health problems associated with aflatoxicosis in Uganda, information about the genetic diversity of the main aflatoxin causing fungus, Aspergillus flavus in this country is still limited. Results A cross-sectional survey was therefore carried out in three main groundnut-growing agro-ecological zones (AEZs) of Uganda; West Nile farming system, Lake Kyoga basin mixed farming system and Lake Victoria basin farming system. This was to assess the genetic diversity of A. flavus and to establish the contamination rates of groundnuts with Aspergillus species at pre- and post-harvest stages. Out of the 213 A. flavus isolates identified in this study, 96 representative isolates were fingerprinted using 16 insertion/deletion microsatellite markers. Data from fingerprinting were analyzed through Neighbor Joining while polymorphism was determined using Arlequin v 3.5. The pre- and post-harvest contamination rates were; 2.5% and 50.0% (West Nile farming system), 55.0% and 35.0% (Lake Kyoga basin mixed farming system) and 32.5% and 32.5% (Lake Victoria basin farming system) respectively. The Chi-square test showed no significant differences between pre- and post-harvest contamination rates among AEZs (p = 0.199). Only 67 out of 96 isolates produced suitable allele scores for genotypic analysis. Analysis of genetic diversity showed higher variation within populations than among populations. Two major clusters (aflatoxigenic and non-aflatoxigenic isolates) were identified as colonizing groundnuts at pre- and postharvest stages. Conclusions These findings provide a first insight on the existence of non-aflatoxigenic strains of A. flavus in Uganda. These strains are potential candidates for developing local Aspergillus biocontrol agent.
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    Stacking three late blight resistance genes from wild species directly into African highland potato varieties confers complete field resistance to local blight races
    (Plant Biotechnology Journal, 2019) Ghislain, Marc; Byarugaba, Arinaitwe Abel; Magembe, Eric; Njoroge, Anne; Rivera, Cristina; Lupe Roman, Marıa; Carlos Tovar, Jose; Gamboa, Soledad; Forbes, Gregory A.; Kreuze, Jan F.; Barekye, Alex; Kiggundu, Andrew
    Considered responsible for one million deaths in Ireland and widespread famine in the European continent during the 1840s, late blight, caused by Phytophthora infestans, remains the most devastating disease of potato (Solanum tuberosum L.) with about 15%–30% annual yield loss in sub-Saharan Africa, affecting mainly smallholder farmers. We show here that the transfer of three resistance (R) genes from wild relatives [RB, Rpi-blb2 from Solanum bulbocastanum and Rpi-vnt1.1 from S. venturii] into potato provided complete resistance in the field over several seasons. We observed that the stacking of the three R genes produced a high frequency of transgenic events with resistance to late blight. In the field, 13 resistant transgenic events with the 3R-gene stack from the potato varieties ‘Desiree’ and ‘Victoria’ grew normally without showing pathogen damage and without any fungicide spray, whereas their non-transgenic equivalent varieties were rapidly killed. Characteristics of the local pathogen population suggest that the resistance to late blight may be long-lasting because it has low diversity, and essentially consists of the single lineage, 2_A1, which expresses the cognate avirulence effector genes. Yields of two transgenic events from ‘Desiree’ and ‘Victoria’ grown without fungicide to reflect small-scale farm holders were estimated to be 29 and 45 t/ha respectively. This represents a three to four-fold increase over the national average. Thus, these late blight resistant potato varieties, which are the farmers’ preferred varieties, could be rapidly adopted and bring significant income to smallholder farmers in sub-Saharan Africa.
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    Variation among banana weevil Cosmopolites sordidus (Germar) populations in Uganda as revealed by AFLP markers and corm damage differences
    (Agriculture & Food Security, 2018) Twesigye, Charles K.; Ssekatawa, Kenneth; Kiggundu, Andrew; Tushemereirwe, Wilberforce; Matovu, Enock
    The banana weevil Cosmopolites sordidus (Germar) is a major production constraint of bananas and plantains (Musa spp.) in the world. Differences in damage levels and pesticide response across regions led to the postulation that there might be considerable variation between banana weevil populations (biotypes) with varying levels of virulence. One of the most sustainable options for banana weevil control is the use of host plant resistance. While new resistant varieties are being developed through both conventional crossbreeding and biotechnology, there is a need to assess the genetic variation of banana weevil populations from eastern, central, southern, southwestern and midwest regions of Uganda to determine whether there are biotypes with different virulence levels. This would help guide new control strategies to target all the possible biotypes. The amplified fragment length polymorphism (AFLP) technique was used to analyze population genetic diversity using four primer combinations (EcoRI/MSeI). Results: Analysis of molecular variance results presented no evidence to support significant genetic variability among the banana weevil populations from eastern, central, southern, southwestern and midwest regions. Practically, all the genetic variation was found to reside within populations (97% for sites and 98% for regions), with only approximately 3% and 2% residing among populations of sites and regions, respectively. Conclusions and recommendations: AFLP markers clustered the banana weevils into two distinct populations consequently supporting the hypothesis of possible presence of banana weevil biotypes in Uganda. However, attempts should be made to make follow-up studies on the seemingly unique population of eastern Uganda using more robust molecular techniques to establish whether the eastern Uganda population constitutes a different biotype.