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Developing wheat varieties with improved ability to use nitrogen (N) efficiently is very desirable,and may offer a sustainable solution to improve crop yields with less fertilizer application.Roots are the main site for nutrient uptake;their size and distribution in soil profiles,and uptake activity largely determine nutrient uptake efficiency.However,at seedling stage,low temperature inhibits root development of winter wheat and nutrient bioavailability in soils;and during grain filling root senescence is not able to meet the increased N demand of high yield potential of modern wheat varieties.Here,we developed transgenic wheat lines with improved NUE by increasing roots ability in acquiring N at seedling stage and N uptake after flowering.We showed that a nitrate-inducible NAC transcription factor TaNAC2-5A could directly bind to the promoter regions of the genes encoding nitrate transporter and glutamine synthetase (GS).Overexpression of TaNAC2-5A in wheat enhanced root growth and nitrate influx rate,and hence increase root ability to acquire nitrogen at seedling stage.Further,we found that TaNAC2-5A-overexpressing transgenic wheat lines had higher grain yield and higher nitrogen accumulation in aerial parts,and allocated more nitrogen in grains in a field experiment.We further analyzed the haplotypes of the plastic GS isoform (GS2) genes in wheat.