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ARS Home » Northeast Area » Washington, D.C. » National Arboretum » Floral and Nursery Plants Research » Research » Publications at this Location » Publication #309326

Research Project: Biotechnology Applied to High Value Ornamental Plants

Location: Floral and Nursery Plants Research

Title: Expression of a synthetic antimicrobial peptide, D4E1, in Gladiolus plants for resistance to Fusarium oxysporum f. sp. gladioli

Author
item Kamo, Kathryn
item Lakshman, Dilip
item Bauchan, Gary
item Rajasekaran, Kanniah - Rajah
item Cary, Jeffrey
item JAYNES, JESSE - Tuskegee University

Submitted to: Plant Cell Tissue and Organ Culture
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 1/11/2015
Publication Date: 1/21/2015
Citation: Kamo, K.K., Lakshman, D.K., Bauchan, G.R., Rajasekaran, K., Cary, J.W., Jaynes, J. 2015. Expression of a synthetic antimicrobial peptide, D4E1, in Gladiolus plants for resistance to Fusarium oxysporum f. sp. gladioli. Plant Cell Tissue And Organ Culture. 121:459-467.

Interpretive Summary: Gladiolus is a popular cutflower and is also grown in gardens. Fusarium oxysporum is the most devastating pathogen of Gladiolus. This soilborne pathogen infects roots resulting in poor plant growth, yellowing of the leaves and death of the plant. It also causes rotting of corms. Previously methyl bromide was used to keep Fusarium under control, but there is no equivalent replacement for it to help the Gladiolus growers. At the USDA, Gladiolus plants were engineered to express an antimicrobial synthetic peptide named D4E1. D4E1 was shown to effectively inhibit growth of pre-germinated Fusarium oxysporum spores. Five lines of Gladiolus that expressed the D4E1 were found to inhibit growth of Fusarium for 10 days, but after a month the plants died from overwhelming proliferation of the fungus.

Technical Abstract: The main pathogen of Gladiolus plants is Fusarium oxysporum, a soilborne fungus that infects roots and corms and kills the plant. Purified D4E1, a synthetic antimicrobial peptide, was found to effectively inhibit 100% of F. oxysporum f. sp. gladioli germinated spores from forming a mycelial mass in vitro at a concentration of 12.5 µM. Gladiolus cv. Peter Pears plants were transformed with the gene for D4E1 using particle bombardment, and 14 independent transformants were obtained. The gene for D4E1 consists of 90 bp and was under the control of the duplicated CaMV 35S promoter. Transgenic plants were tested in vitro for resistance to F. oxysporum, and several lines appeared to be more resistant than the control plants that lacked D4E1. Cell extracts of transgenic Gladiolus lines 6(1) and 7(1) inhibited growth of F. oxysporum f. sp. gladioli germinated spores by 34 and 38%, respectively, in vitro. F. oxysporum f. sp. gladioli was transformed with the ECFP (cyan) gene allowing us to follow the growth of F. oxysporum during infection of D4E1-transformed and non-transformed roots. Fluorescence observations using a confocal laser scanning microscope showed that 3-10 days after infection, F. oxysporum covered the surface of the root and formed pseudo-appressoria, but hyphae were never observed to penetrate cells of the root. Ten days after infection with F. oxysporum, non-transformed roots had completely disintegrated whereas transgenic roots of line 7(1) were just beginning to lose their cellular integrity.