An efficient mannose selection protocol for tomato that has no adverse effect on the ploidy level of transgenic plants.
A protocol for Agrobacterium-mediated transformation with mannose selection was developed for cotyledon petiole, hypocotyl and leaf explants of tomato (Lycopersicon esculentum L. Mill). More than 400 transgenic plants from three tomato varieties were selected with 1% mannose in combination with 0.1-0.5% glucose. Average transformation frequencies ranged from 2.0 to 15.5% depending on the construct, genotype and type of tissue used for transformation. The highest transformation rate was obtained for hypocotyl explants from tomato variety SG048. The ploidy levels of 264 independent transgenic events and 233 non-transgenic plants regenerated from tissue culture were assessed by flow cytometry. The incidence of polyploids within the total population of transgenic plants varied from 10 to 78% and was not significantly different from the non-transgenic population. The greatest variation in the proportion of polyploids was observed in plants derived from different explant types, both in transgenic and non-transgenic regenerants, across three studied genotypes. Transgenic and non-transgenic plants regenerated from leaves included the highest number of normal diploid plants (82-100%), followed by cotyledon petiole-derived plants (63-78%). Transgenic plants produced from hypocotyls contained 22-58% diploids depending on the genotype used in transformation. Results described in this study demonstrate that, although transformation frequencies for leaf tissue are still lower under current protocols, the high percentage of diploids obtained make leaf tissue an attractive transformation target.
Sigareva
Marina
M
Syngenta Biotechnology, 3054 Cornwallis Road, Research Triangle Park, NC 27709-2257, USA. marina.sigareva@syngenta.com
Spivey
Rody
R
Willits
Michael G
MG
Kramer
Catherine M
CM
Chang
Yin-Fu
YF
D016960:Agrobacterium tumefaciens / Q000235:genetics; D018744:DNA, Plant / Q000032:analysis / Q000235:genetics; D018628:Gene Dosage; D018506:Gene Expression Regulation, Plant / Q000235:genetics; D005822:Genetic Vectors / Q000235:genetics; D005838:Genotype; D018546:Hypocotyl / Q000235:genetics / Q000254:growth & development / Q000378:metabolism; D018551:Solanum lycopersicum / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D008358:Mannose / Q000096:biosynthesis / Q000235:genetics*; D030821:Plants, Genetically Modified / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D011003:Ploidies; D013045:Species Specificity; D046509:Tissue Culture Techniques / Q000379:methods; D014162:Transfection / Q000379:methods; D014170:Transformation, Genetic / Q000235:genetics*; D019076:Transgenes / Q000235:genetics
D018744:DNA, Plant; D008358:Mannose