Genetic transformation of garlic (Allium sativum L.) with tobacco chitinase and glucanase genes for tolerance to the fungus Sclerotium cepivorum

Abstract

Garlic yield and quality have decreased due to white rot disease caused by Sclerotium cepivorum Berk. A transformation protocol to introduce tobacco chitinase and glucanase genes into garlic embryogenic calli using Agrobacterium tumefaciens has been established. LBA4404 strain having pC2301CHGLU plasmid with TaCh, glu, gus and nptII genes (coding for chitinase, glucanase, β-glucuronidase and neomycin phosphotransferase, respectively) was used. 30 putative transgenic clones were obtained from inoculated calli after six months. Histochemical assay revealed high gus activity in 43% of the clones. Molecular analysis of transgenic plants showed 92% of the clones carried TaCh gene. Eight culture media for plant regeneration from transgenic calli were evaluated; MTDZ-1 (thidiazuron 1 mg/l) medium induced the highest number of plants (38.4 plants). Transgenic plants were grown in the greenhouse and they developed normally. S. cepivorum in vitro bioassays showed 41 to 60% of mycelial invasion in the transgenic plants, and 80% in non-transgenic plants (control). Transformed plants were not completely resistant, but they showed a delay in fungal infection. This is the first report on the introduction of fungal resistance genes in garlic.

Keywords: Allium sativum, Agrobacterium tumefaciens, Sclerotium cepivorum, glucanase, chitinase

African Journal of Biotechnology Vol. 12(22), pp. 3482-3492