A simple shoot multiplication procedure using internode explants, and its application for particle bombardment and Agrobacterium-mediated transformation in watercress.
A shoot multiplication system derived from internode explants was investigated with the aim of improving genetic characteristics of watercress (Nasturtium officinale R. Br.). Internodes of ca. 1 cm excised from in vitro stock shoot culture were placed on half-strength Murashige and Skoog (MS) medium supplemented with 3 muM 2,4-dichlorophenoxyacetic acid as a pre-treatment. Laser scanning microscopy indicated clearly that the first sign of meristematic cell division could be seen after 1-2 days of pre-culture, and meristematic tissues multiplied along the vascular cambium of the internode segment during 7 days of culture. Multiple shoots could be obtained from more than 90% of the pre-treated explants when they were subsequently transferred to MS medium supplemented with 1 muM thidiazuron for 3 weeks. These findings indicate that pre-treatment of the internodes for 7 days promoted their capacity for organogenesis. Using this pre-treatment, frequent generation of transgenic watercress plants was achieved by adapting particle bombardment and Agrobacterium-mediated transformation techniques with a construct expressing a synthetic green florescent protein gene.
Ogita
Shinjiro
S
Laboratory of Plant and Cell Engineering, Department of Biotechnology, Toyama Prefectural University, Imizu, Toyama 939-0398, Japan. ogita@pu-toyama.ac.jp
Usui
Miki
M
Shibutani
Nanae
N
Kato
Yasuo
Y
D019470:Biolistics / Q000379:methods*; D019607:Brassicaceae / Q000166:cytology / Q000235:genetics* / Q000382:microbiology*; D020869:Gene Expression Profiling; D049452:Green Fluorescent Proteins / Q000235:genetics; D019697:Onions / Q000235:genetics; D038081:Organogenesis / Q000187:drug effects; D010937:Plant Growth Regulators / Q000494:pharmacology; D018520:Plant Shoots / Q000166:cytology / Q000187:drug effects / Q000196:embryology / Q000254:growth & development*; D018547:Plant Stems / Q000166:cytology / Q000187:drug effects / Q000196:embryology / Q000254:growth & development*; D030821:Plants, Genetically Modified; D012231:Rhizobium / Q000378:metabolism*; D014170:Transformation, Genetic / Q000187:drug effects
D010937:Plant Growth Regulators; D049452:Green Fluorescent Proteins