Arabidopsis and Musa cyclin D2 expressed in banana (cv. “Sukali Ndiizi”- AAB) enhances regeneration efficiency

Genetic transformation of banana is important because of its polyploidy, sterility and long generation time of most cultivars which limit conventional breeding. However, transformability and regeneration of transgenic lines remains low in bananas. This research reports on the potential of CycD2 genes to improve transformation and regeneration efficiency of banana (cv. “Sukali Ndiizi”). Two genes Arath;CycD2;1 and Musa;CycD2;1 were evaluated for cell cycle modification of the embryogenic cell suspension that is conventionally used in banana genetic engineering at the National Biotechnology Centre, Kawanda. The UidA (GUS) gene was used as reporter gene to establish transient transformation frequency by fusing it with each of the CycD2; 1 genes and Cauliflower mosaic virus 35S promoter in the binary vector, pC1305.1. The transformed “Sukali Ndiizi” cells were cultured on selection media and the hygromycin resistant clones developed into shoots. The Gus assay analyses showed a success rate of 80 to 90% for all the constructs including the control, transformed with the empty vector without CycD2; 1 gene. Also, the Gus assay of the regenerants showed that the gene was expressed in different parts of the plants (roots, corm and leaves). Polymerase chain reaction (PCR) analysis of the regenerated shoots gave the regeneration frequency of the embryogenic clones of Arath; CycD2; 1 and Musa; CycD2; 1 gene was 47 and 62%, respectively. This was much higher than that of the control clones without CycD2; 1 (18%). The results show that CycD2; 1 genes have the potential to significantly improve regeneration efficiency of “Sukali Ndiizi” cells”.Keywords: Cell cycle genes, reporter gene, genetic transformation, regeneration efficiencyAfrican Journal of Biotechnology Vol. 12(13), pp. 1467-147

DETERMINANTS OF RESOURCE ALLOCATION IN LOW INPUT AGRICULTURE: THE CASE OF BANANA PRODUCTION IN UGANDA

Replaced with revised version of paper 09/01/04.Crop Production/Industries,

Building a bi-directional promoter binary vector from the intergenic region of Arabidopsis thaliana cab1 and cab2 divergent genes useful for plant transformation

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.Key words: Arabidopsis cab genes, bidirectional promoters, gene expression, plant genetic engineering, reporter genes, photosynthetic genes

Enhancing banana weevil (Cosmopolites sordidus) resistance by plant genetic modification: A perspective

Banana weevil is a serious pest of bananas and plantains in Africa. The development of resistant cultivars is seen as the long term and more sustainable control strategy. The difficulty in conventional breeding of bananas and plantains has prompted efforts towards the use of genetic transformation for banana and plantain improvement. In this review, the current status of banana weevil resistance, sources of resistance and resistance mechanisms is assessed. Further, current efforts and future prospects for identifying resistance genes outside the genus Musa with potential to control banana weevil in a transgenic approach are outlined and discussed. Key words: Banana weevil, host plant resistance, pest resistance genes, transgenic plants African Journal of Biotechnology Vol. 2 (12), pp. 563-569, December 200

Inhibition of cell death as an approach for development of transgenic resistance against Fusarium wilt disease

Fusarium oxysporum f. sp. cubense (Foc) is one of the major threats to dessert banana (Musa spp.) production. In Uganda, ‘Sukali Ndiizi’ is one of the most popular dessert banana cultivars and it is highly susceptible to Fusarium wilt. Development of resistant cultivars through transgenic approaches has shown to offer one of the most effective control options for most diseases. The transgenic approaches for providing plant disease resistance have mainly been through either enzymatic destruction of pathogen structures, neutralization of pathogen and its products or production of metabolites that eventually kill the pathogen. However in recent years, methods that prevent cell death of host plant after infection especially for necrotrophic pathogens like F. oxysporum have registered success in providing resistance in several crops. We investigated whether the transgenic expression of a programmed cell death inhibition gene in Sukali Ndiizi could be used to confer Fusarium resistance to Foc race 1. Embryogenic cell suspensions of cv. ‘Sukali Ndiizi, were stably transformed with a synthetic, plant-codon optimise mCed-9 gene. Twenty-eight independently transformed plant lines were regenerated. The lines were inoculated with Foc race 1 and observed for 13 weeks in small-plant glasshouse. Three transgenic lines showed significantly lower internal and external disease symptoms than the wild-type susceptible ‘Sukali Ndiizi’ banana plants used as controls. This is the first report from Africa on the generation of Fusarium wilt tolerant transgenic ‘Sukali Ndiizi’, a very popular but rapidly diminishing African dessert banana.Key words: Fusarium wilt, banana, Sukali Ndiizi, Fusarium oxysporum f. sp. Cubense race 1, programmed cell death, disease resistance

Agrobacterium tumefaciens-mediated transformation of Pseudocercospora fijiensis to determine the role of PfHog1 in osmotic stress regulation and virulence modulation

Open Access Journal; Published online: 16 May 2017Black Sigatoka disease, caused by Pseudocercospora fijiensis is a serious constraint to banana production worldwide. The disease continues to spread in new ecological niches and there is an urgent need to develop strategies for its control. The high osmolarity glycerol (HOG) pathway in Saccharomyces cerevisiae is well known to respond to changes in external osmolarity. HOG pathway activation leads to phosphorylation, activation and nuclear transduction of the HOG1 mitogen-activated protein kinases (MAPKs). The activated HOG1 triggers several responses to osmotic stress, including up or down regulation of different genes, regulation of protein translation, adjustments to cell cycle progression and synthesis of osmolyte glycerol. This study investigated the role of the MAPK-encoding PfHog1 gene on osmotic stress adaptation and virulence of P. fijiensis. RNA interference-mediated gene silencing of PfHog1 significantly suppressed growth of P. fijiensis on potato dextrose agar media supplemented with 1 M NaCl, indicating that PfHog1 regulates osmotic stress. In addition, virulence of the PfHog1-silenced mutants of P. fijiensis on banana was significantly reduced, as observed from the low rates of necrosis and disease development on the infected leaves. Staining with lacto phenol cotton blue further confirmed the impaired mycelial growth of the PfHog1 in the infected leaf tissues, which was further confirmed with quantification of the fungal biomass using absolute- quantitative PCR. Collectively, these findings demonstrate that PfHog1 plays a critical role in osmotic stress regulation and virulence of P. fijiensis on its host banana. Thus, PfHog1 could be an interesting target for the control of black Sigatoka disease in banana

Silencing of the mitogen-activated protein kinases (MAPK) fus3 and slt2 in pseudocercospora fijiensis reduces growth and virulence on host plants

Open Access JournalPseudocercospora fijiensis, causal agent of the black Sigatoka disease (BSD) of Musa spp., has spread globally since its discovery in Fiji 1963 to all the banana and plantain growing areas across the globe. It is becoming the most damaging and economically important disease of this crop. The identification and characterization of genes that regulate infection processes and pathogenicity in P. fijiensis will provide important knowledge for the development of disease-resistant cultivars. In many fungal plant pathogens, the Fus3 and Slt2 are reported to be essential for pathogenicity. Fus3 regulates filamentous-invasion pathways including the formation of infection structures, sporulation, virulence, and invasive and filamentous growth, whereas Slt2 is involved in the cell-wall integrity pathway, virulence, invasive growth, and colonization in host tissues. Here, we used RNAi-mediated gene silencing to investigate the role of the Slt2 and Fus3 homologs in P. fijiensis in pathogen invasiveness, growth and pathogenicity. The PfSlt2 and PfFus3 silenced P. fijiensis transformants showed significantly lower gene expression and reduced virulence, invasive growth, and lower biomass in infected leaf tissues of East African Highland Banana (EAHB). This study suggests that Slt2 and Fus3 MAPK signaling pathways play important roles in plant infection and pathogenic growth of fungal pathogens. The silencing of these vital fungal genes through host-induced gene silencing (HIG) could be an alternative strategy for developing transgenic banana and plantain resistant to BSD

First report of banana bunchy top disease caused by Banana bunchy top virus in Uganda

Banana bunchy top virus (BBTV) that causes banana bunchy top disease (BBTD) is ranked among the top 100 invasive species in the world. Despite being omnipresent in the Democratic Republic of Congo (DR Congo) and South-Western Rwanda for over 2 decades, BBTV had not been reported in neighbouring Uganda. However, in 2020, banana plants with BBTD characteristic symptoms were observed in Arua City located in the North Western part of Uganda, bordering DR Congo. BBTV in these plants was confirmed using PCR and the comparison of the genomic DNA nucleotide sequence with nucleotide sequences in NCBI data base (https://www.ncbi.nlm.nih.gov/). The gDNA nucleotide sequence had 98-99% similarity with BBTV isolates form different countries in Africa, Asia, South Pacific and the USA. These results constitute the first confirmation of BBTD in Uganda. This infection is anticipated to have been introduced from DR Congo, mainly through infected planting materials. Once established, BBTD is very difficult to control on small-scale farms. It is therefore crucial to urgently understand the current spread of the virus, determine its risk to banana production in Uganda and devise proactive measures for its management

'NABIO808' (Syn. 'NAROBAN5'): A tasty cooking banana cultivar with resistance to pests and diseases

Abstract 'NABIO808' is a newly released, conventionally-bred triploid cooking banana cultivar in Uganda. It produces an average bunch yield of 54.5 t ha-1 yr-1 and is resistant to weevils, nematodes, and black Sigatoka. Additionally, its food is yellow, soft, and tasty, like that of most preferred landrace cultivars, making it more acceptable to end-users