Banana Xanthomonas wilt (BXW), caused by Xanthomonas campestris pv. musacearum, is one of the most important diseases of banana (Musa sp.) and currently considered as the biggest threat to banana production in Great Lakes region of East and Central Africa. The pathogen is highly contagious and its spread has endangered the livelihood of millions of farmers who rely on banana for food and income. The development of disease resistant banana cultivars remains a high priority since farmers are reluctant to employ labor-intensive disease control measures and there is no host plant resistance among banana cultivars. In this study, we demonstrate that BXW can be efficiently controlled using transgenic technology. Transgenic bananas expressing the plant ferredoxin-like protein (Pflp) gene under the regulation of the constitutive CaMV35S promoter were generated using embryogenic cell suspensions of banana. These transgenic lines were characterized by molecular analysis. After challenge with X. campestris pv. musacearum transgenic lines showed high resistance. About 67% of transgenic lines evaluated were completely resistant to BXW. These transgenic lines did not show any disease symptoms after artificial inoculation of in vitro plants under laboratory conditions as well as potted plants in the screen-house, whereas non-transgenic control plants showed severe symptoms resulting in complete wilting. This study confirms that expression of the Pflp gene in banana results in enhanced resistance to BXW. This transgenic technology can provide a timely solution to the BXW pandemic.

Application of biotechnological tools in breeding, germplasm conservation and propagation of the East African bananas (Musa sp.) is limited by the crop’s recalcitrance to somatic embryogenesis. This study was undertaken to establish an efficient callus induction and plant regeneration protocol from immature male flowers in the most commercial and farmer preferred banana cultivar “Nakitembe”. Embryogenic callusing was improved from 2.9% on conventional MA1 culture medium to 4.2% with 500 mg L-1 L-Glutamine, 500 mg L-1 L-Proline and 300 mg L-1 casein hydrolysate supplements. A combination of the three amino acids with Chui N6 salts yielded the highest callusing of 10.2%. Cell suspensions were developed and maintained on conventional MS based MA2 medium. Subsequently, the highest efficiency of embryos germination (up to 80%) was achieved on MA4 medium, supplemented with 2.25 mg L-1 BAP and 0.2 mg L-1 IAA. The developed protocol has been successfully applied in Agrobacterium mediated genetic transformations of this variety.

Gene transfer into the plant cell is a key step towards its successful genetic modification and its efficiency is heavily dependent on plant and bacterial cell biological status and a wide array of physical conditions. Gene transfer efficiencies in East African Highland Banana (EAHB) cell lines remains low compared to other monocotyledonous crops like rice and wheat due to factors such as high oxidative stress. The use of antioxidants is fundamental in influencing gene transfer events during Agrobacterium-plant cell co-cultivation. Here we report significant enhancement of gene transfer efficiency in the EAHB cultivar 'Nakinyika' (EA-AAA) by supplementing co-cultivation medium with antioxidants; ascorbic acid (AA), glutathione (GSH), tocopherol (TOC) and silver nitrate (SN). The most enhancing antioxidant by number of blue foci after histochemical assay, as a parameter of gene transfer efficiency, was ascorbic acid (174 cells) at a concentration of 40 mg/L, followed by glutathione (91 cells) and tocopherol (91 cells) both at 50 mg/L. The least enhancement was observed when ascorbic acid (39 cells), silver nitrate (41 and 31 cells) were used at concentrations of 20, 4 and 6 mg/L compared to 72 cells in controls (no anti-oxidants). Regeneration efficiency increased from 29 cells in controls to 46% for SN at 8 mg/L; 43% for GSH at 100 mg/L; 30% for TOC at 75 mg/L; and 48% for AA at 20 mg/L. Polymerase chain reaction (PCR) results using gusA specific primers showed that these regenerants were putative transformants and grew normally during regeneration, rooting and multiplication. GSH and TOC significantly enhanced gene transfer efficiency while AA and SN showed significant increases in shoot regeneration compared to controls. The current results show that antioxidants significantly enhance gene transfer and regeneration efficiency in recalcitrant banana cell lines and could significantly enhance the overall transformation efficiencies in cases where numerous transgenic lines are required in a short time.