A novel mannose-based selection system for plant transformation using celery mannose-6-phosphate reductase gene.
To investigate its potential application as a selectable marker for plant transformation, the mannitol producing, celery mannose-6-phosphate reductase gene (M6PR) was transformed into Arabidopsis and tobacco using Agrobacterium tumefaciens-mediated transformation. Mannose-tolerance assays in transgenic materials revealed that the M6PR can act as a selectable marker gene in either a positive or a negative selection mode depending on the plant species. For mannose sensitive species, such as Arabidopsis, expression of M6PR enhanced mannose tolerance and provided a positive selection for transgenic seeds. On medium containing 2 g/L mannose, transgenic seeds germinated, whereas wild type (WT) seeds did not. For mannose-tolerant species, expression of M6PR increased mannose sensitivity in tobacco and enabled a negative selection for transgenic leaves and seeds. Mannose at 30 g/L blanched leaf explants from all 29 transgenic tobacco events with M6PR. In contrast, 30 g/L mannose did not inhibit shoot regeneration from leaf explants of WT or transgenic plants with either an antisense M6PR or a plasmid control. Similarly, mannose at 30 g/L inhibited seed germination of transgenic tobacco seeds with M6PR but not that of WT or transgenic tobacco with either the antisense M6PR or the plasmid control. Northern blot confirmed transcripts of the M6PR in transgenic tobacco, and accumulation of mannitol verified activity of the M6PR in tobacco leaves. Either positive or negative selection using the celery M6PR is versatile for plant transformation. Additionally, the celery M6PR is a potential target gene for improving salt-tolerance in plants due to mannitol accumulation.
Song
Guo-Qing
GQ
Plant Biotechnology Resource and Outreach Center, Department of Horticulture, Michigan State University, East Lansing, MI 48824, USA.
Sink
Kenneth C
KC
Ma
Yumin
Y
Herlache
Thomas
T
Hancock
James F
JF
Loescher
Wayne H
WH
D016960:Agrobacterium tumefaciens / Q000235:genetics; D000077102:Apium / Q000201:enzymology*; D017360:Arabidopsis / Q000235:genetics* / Q000378:metabolism; D018506:Gene Expression Regulation, Plant; D005818:Genetic Engineering / Q000379:methods*; D018525:Germination; D008353:Mannitol / Q000378:metabolism; D008358:Mannose / Q000378:metabolism; D018515:Plant Leaves / Q000235:genetics / Q000378:metabolism; D030821:Plants, Genetically Modified / Q000235:genetics / Q000378:metabolism; D012639:Seeds / Q000235:genetics / Q000378:metabolism; D013401:Sugar Alcohol Dehydrogenases / Q000235:genetics*; D014026:Nicotiana / Q000235:genetics* / Q000378:metabolism; D014170:Transformation, Genetic
D008353:Mannitol; D013401:Sugar Alcohol Dehydrogenases; C076442:mannose-6-phosphate reductase; D008358:Mannose