Agrobacterium-mediated transformation of Eruca sativa

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Journal titlePlant Cell, Tissue and Organ Culture
Pages253260; # of pages: 8
Subject6-benzyladenine; Acetyl transferase; Agrobacterium mediated transformations; Agrobacterium tumefaciens; Agrobacterium tumefaciens-mediated transformation; Bacterial cells; Brassicaceae; Eruca sativa; Fusion genes; Hypocotyl explants; Indole-3-acetic acid; Kanamycins; Naphthaleneacetic acid; Phosphinothricin; Putative transformants; Regenerants; Silver nitrates; Southern blot analysis; Trans-genes; Transformation frequency; Transgene; Transgenic lines; Acetic acid; Herbicides; Plants (botany); Genes; Agrobacterium; Agrobacterium tumefaciens; Bacteria (microorganisms); Brassicaceae; Eruca; Eruca sativa; Rhizobium
AbstractAn Agrobacterium tumefaciens-mediated transformation system was developed for Eruca sativa (eruca). Hypocotyl explants were co-cultivated with bacterial cells carrying a plasmid harboring a uidA:nptII fusion gene along a phosphinothricin acetyl transferase (PAT) gene cassette, for a period of 2 days. These were grown on a high cytokinin/auxin medium containing 5.0 mg l -1 6-benzyladenine (BA), 1.0 mg l -1 indole-3-acetic acid (IAA), and 0.1 mg l -1 α-naphthaleneacetic acid (NAA). Explants were then transferred to a lower cytokinin/auxin medium containing 2.0 mg l -1 BA and 0.1 mg l -1 NAA along with 5.0 mg l -1 silver nitrate and 300 mg l -1 Timentin®. Upon transfer to a selection medium containing either 20 mg l -1 kanamycin or 2 mg l -1 L-phosphinothricin (L-ppt), shoot regenerants were observed. Expression of the transgenes in putative transformants was confirmed using a histochemical GUS assay. Presence of the PAT transgene in GUS-positive T0 plants was confirmed by Southern blot analysis. Moreover, spot tests of T1 seedlings were conducted using the L-ppt herbicide. A transformation frequency of 1. 1% was obtained with more than 60% of transgenic lines containing single copies of the transgenes. © 2011 Springer Science+Business Media B.V.
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AffiliationNational Research Council Canada (NRC-CNRC); NRC Plant Biotechnology Institute (PBI-IBP)
Peer reviewedYes
NPARC number21271217
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Record identifierf85d8d85-e53d-43b4-b89f-33432334155a
Record created2014-03-24
Record modified2016-05-09
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