title: Enhanced resistance to Sclerotinia sclerotiorum in Brassica napus by co-expression of defensin and chimeric chitinase genes.
abstract: Sclerotinia stem rot caused by Sclerotinia sclerotiorum is one of the major fungal diseases of Brassica napus L. To develop resistance against this fungal disease, the defensin gene from Raphanus sativus and chimeric chit42 from Trichoderma atroviride with a C-terminal fused chitin-binding domain from Serratia marcescens were co-expressed in canola via Agrobacterium-mediated transformation. Twenty transformants were confirmed to carry the two transgenes as detected by polymerase chain reaction (PCR), with 4.8 % transformation efficiency. The chitinase activity of PCR-positive transgenic plants were measured in the presence of colloidal chitin, and five transgenic lines showing the highest chitinase activity were selected for checking the copy number of the transgenes through Southern blot hybridisation. Two plants carried a single copy of the transgenes, while the remainder carried either two or three copies of the transgenes. The antifungal activity of two transgenic lines that carried a single copy of the transgenes (T4 and T10) was studied by a radial diffusion assay. It was observed that the constitutive expression of these transgenes in the T4 and T10 transgenic lines suppressed the growth of S. sclerotiorum by 49 % and 47 %, respectively. The two transgenic lines were then let to self-pollinate to produce the T2generation. Greenhouse bioassays were performed on the transgenic T2young leaves by challenging with S. sclerotiorum and the results revealed that the expression of defensin and chimeric chitinase from a heterologous source in canola demonstrated enhanced resistance against sclerotinia stem rot disease.
authors: National Institute of Genetic Engineering and Biotechnology (NIGEB), Tehran, Iran.
authors: Nasim
authors: N
authors: Zarinpanjeh
authors: National Institute of Genetic Engineering and Biotechnology (NIGEB), Tehran, Iran. motalebi@nigeb.ac.ir.
authors: Mostafa
authors: M
authors: Motallebi
authors: National Institute of Genetic Engineering and Biotechnology (NIGEB), Tehran, Iran.
authors: Mohammad Reza
authors: MR
authors: Zamani
authors: National Institute of Genetic Engineering and Biotechnology (NIGEB), Tehran, Iran.
authors: Mahboobeh
authors: M
authors: Ziaei
mesh_terms: D060054:Agrobacterium; D001203:Ascomycota; D029688:Brassica napus / Q000235:genetics* / Q000382:microbiology; D002688:Chitinases / Q000235:genetics*; D023082:Defensins / Q000235:genetics*; D060467:Disease Resistance / Q000235:genetics*; D009842:Oligopeptides; D010935:Plant Diseases / Q000235:genetics* / Q000382:microbiology; D030821:Plants, Genetically Modified / Q000235:genetics / Q000382:microbiology; D031224:Raphanus / Q000235:genetics; D014170:Transformation, Genetic; D014242:Trichoderma / Q000201:enzymology / Q000235:genetics
chemical_list: C570180:AFP1 peptide, Trapa natans; D023082:Defensins; D009842:Oligopeptides; D002688:Chitinases
keywords: Antifungal activity; Brassica napus; Chimeric chitinase; Defensin; Sclerotinia sclerotiorum