title: Retransformation of marker-free potato for enhanced resistance against fungal pathogens by pyramiding chitinase and wasabi defensin genes.
abstract: Multi-auto-transformation vector system has been one of the strategies to produce marker-free transgenic plants without using selective chemicals and plant growth regulators and thus facilitating transgene stacking. In the study reported here, retransformation was carried out in marker-free transgenic potato CV. May Queen containing ChiC gene (isolated from Streptomyces griseus strain HUT 6037) with wasabi defensin (WD) gene (isolated from Wasabia japonica) to pyramid the two disease resistant genes. Molecular analyses of the developed shoots confirmed the existence of both the genes of interest (ChiC and WD) in transgenic plants. Co-expression of the genes was confirmed by RT-PCR, northern blot, and western blot analyses. Disease resistance assay of in vitro plants showed that the transgenic lines co-expressing both the ChiC and WD genes had higher resistance against the fungal pathogens, Fusarium oxysporum (Fusarium wilt) and Alternaria solani (early blight) compared to the non-transformed control and the transgenic lines expressing either of the ChiC or WD genes. The disease resistance potential of the transgenic plants could be increased by transgene stacking or multiple transformations.
authors.affiliation: Laboratory of Plant Cell Technology, Graduate School of Horticulture, Chiba University, 648 Matsudo, Matsudo, Chiba, 271-8510, Japan, rahamsher75@hotmail.com.
authors.forename: Raham Sher
authors.initials: RS
authors.lastname: Khan
authors.forename: Nader Ahmed
authors.initials: NA
authors.lastname: Darwish
authors.forename: Bushra
authors.initials: B
authors.lastname: Khattak
authors.forename: Valentine Otang
authors.initials: VO
authors.lastname: Ntui
authors.forename: Kynet
authors.initials: K
authors.lastname: Kong
authors.forename: Kazuki
authors.initials: K
authors.lastname: Shimomae
authors.forename: Ikuo
authors.initials: I
authors.lastname: Nakamura
authors.forename: Masahiro
authors.initials: M
authors.lastname: Mii
mesh_terms: D000528:Alternaria / Q000472:pathogenicity*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D002688:Chitinases / Q000235:genetics / Q000378:metabolism*; D023082:Defensins / Q000235:genetics / Q000378:metabolism*; D005670:Fusarium / Q000472:pathogenicity*; D066298:In Vitro Techniques; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism; D018520:Plant Shoots / Q000378:metabolism; D030821:Plants, Genetically Modified / Q000235:genetics / Q000382:microbiology*; D011198:Solanum tuberosum / Q000235:genetics* / Q000382:microbiology; D013305:Streptomyces griseus / Q000201:enzymology; D014170:Transformation, Genetic; D031228:Wasabia / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D023082:Defensins; D010940:Plant Proteins; D002688:Chitinases