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GrainGenes Reference Report: PRT-20-331

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Reference	PRT-20-331
Title	Visual screening of microspore-derived transgenic barley (Hordeum vulgare L.) with green-fluorescent protein
Journal	Plant Cell Reports
Year	2001
Volume	20
Pages	331-337
Author	Carlson AR Letarte J Chen J Kasha KJ
Abstract	Summary: The green-fluorescent protein (GFP) gene from the Pacific Northwest jellyfish, Aequorea victoria, was used as a screenable marker in the production of transgenic barley plants. Isolated barley microspore culture was biolistically transformed with two synthetic forms of GFP, sgfp and pgfp. Thirty-seven fluorescing multicellular structures were isolated using epifluorescent microscopy. Sixteen structures developed shoots, but only five regenerated into green plants. Three events had been co-bombarded with beta-glucuronidase (gus) and assayed positive for gus expression in the leaves, and all five events were positive for gfp expression. The expected transgene band size was PCR-amplified from all five plants, and Southern blots performed on three plants revealed unique patterns of gfp transgene integration. Fluorescent in situ hybridization also revealed the transgenic status and hemizygous nature of all the events. GFP-based visual screening provides a viable alternative method to chemical selection of transgenic plants from barley microspore culture
Keyword	[ Hide all but 1 of 46 ] barley barley microspore culture barley plants barley transformation biolistic transformation culture events fluorescent in situ hybridization gene gfp green fluorescent protein gus gus expression hordeum hordeum vulgare hybridization in situ in situ hybridization integration jellyfish marker method microscopy microspore microspore culture particle bombardment pattern plant transformation protein regeneration reporter screenable marker screening selection shoot situ hybridization structure transgene transgene integration transgenic transgenic barley transgenic barley plants transgenic plant transient victoria visual screening

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