title: Improvement ofAgrobacterium-mediated transformation of cucumber (Cucumis sativusL.) by combination of vacuum infiltration and co-cultivation on filter paper wicks.
abstract: An improved method for genetic transformation of cucumber (Cucumis sativusL. cv. Shinhokusei No. 1) was developed. Vacuum infiltration of cotyledonary explants withAgrobacteriumsuspension enhanced the efficiency ofAgrobacteriuminfection in the proximal regions of explants. Co-cultivation on filter paper wicks suppressed necrosis of explants, leading to increased regeneration efficiency. Putative transgenic plants were screened by kanamycin resistance and green fluorescent protein (GFP) fluorescence, and integration of the transgene into the cucumber genome was confirmed by genomic polymerase chain reaction (PCR) and Southern blotting. These transgenic plants grew normally and T1seeds were obtained from 7 lines. Finally, stable integration and transmission of the transgene in T1generations were confirmed by GFP fluorescence and genomic PCR. The average transgenic efficiency for producing cucumbers with our method was 11.9 ± 3.5 %, which is among the highest values reported until date using kanamycin as a selective agent.
authors.affiliation: Genetically Modified Organism Research Center, National Institute of Agrobiological Sciences, 2-1-2 Kannondai, Tsukuba, Ibaraki 305-8602 Japan.
authors.forename: Yoshihiko
authors.initials: Y
authors.lastname: Nanasato
authors.forename: Ken-Ichi
authors.initials: KI
authors.lastname: Konagaya
authors.forename: Ayako
authors.initials: A
authors.lastname: Okuzaki
authors.forename: Mai
authors.initials: M
authors.lastname: Tsuda
authors.forename: Yutaka
authors.initials: Y
authors.lastname: Tabei
keywords: Acetosyringone; Agrobacterium; Cucumis sativus; Filter paper wicks; Transformation; Vacuum infiltration