Targeted modification of storage protein content resulting in improved amino acid composition of barley grain.
C-hordein in barley and ω-gliadins in wheat are members of the prolamins protein families. Prolamins are the major component of cereal storage proteins and composed of non-essential amino acids (AA) such as proline and glutamine therefore have low nutritional value. Using double stranded RNAi silencing technology directed towards C-hordein we obtained transgenic barley lines with up to 94.7% reduction in the levels of C-hordein protein relative to the parental line. The composition of the prolamin fraction of the barley parental line cv. Golden Promise was resolved using SDS-PAGE electrophoresis, the protein band were excised and the proteins identified by quadrupole-time-of-flight mass spectrometry. Subsequent SDS-PAGE separation and analysis of the prolamin fraction of the transgenic lines revealed a reduction in the amounts of C-hordeins and increases in the content of other hordein family members. Analysis of the AA composition of the transgenic lines showed that the level of essential amino acids increased with a concomitant reduction in proline and glutamine. Both the barley C-hordein and wheat ω-gliadin genes proved successful for RNAi-gene mediated suppression of barley C-hordein level. All transgenic lines that exhibited a reduction for C-hordein showed off-target effects: the lines exhibited increased level of B/γ-hordein while D-hordein level was reduced. Furthermore, the multicopy insertions correlated negatively with silencing.
Science and Technology, Department of Molecular Biology and Genetics, Research Centre Flakkebjerg, Aarhus University, 4200, Slagelse, Denmark.|Department of Agronomy, Hajee Mohammad Danesh Science and Technology University, Dinajpur, 5200, Bangladesh.
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Sikdar
Verzyme (UK) Ltd., Plas Gogerddan, Aberystwyth, Wales, SY23 3EB, UK.
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Bowra
Science and Technology, Department of Molecular Biology and Genetics, Research Centre Flakkebjerg, Aarhus University, 4200, Slagelse, Denmark.|Departamento de Genética, Escola Superior de Agricultura Luiz de Queiroz, Universidade de São Paulo, Piracicaba, SP, CEP 13418-900, Brazil.
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Schmidt
Science and Technology, Department of Molecular Biology and Genetics, Research Centre Flakkebjerg, Aarhus University, 4200, Slagelse, Denmark.
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Dionisio
Science and Technology, Department of Molecular Biology and Genetics, Research Centre Flakkebjerg, Aarhus University, 4200, Slagelse, Denmark.
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Holm
Science and Technology, Department of Molecular Biology and Genetics, Research Centre Flakkebjerg, Aarhus University, 4200, Slagelse, Denmark. eva.vincze@mbg.au.dk.
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Vincze
D000596:Amino Acids / Q000737:chemistry* / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel; D005903:Gliadin / Q000235:genetics; D005983:Glutens / Q000235:genetics* / Q000378:metabolism; D001467:Hordeum / Q000737:chemistry* / Q000235:genetics*; D030821:Plants, Genetically Modified; D055315:Prolamins / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D034622:RNA Interference; D012639:Seeds / Q000737:chemistry* / Q000235:genetics; D053719:Tandem Mass Spectrometry / Q000379:methods; D014908:Triticum / Q000235:genetics
D000596:Amino Acids; D055315:Prolamins; D005983:Glutens; D005903:Gliadin
Alteration multigene families; Antisense transgenic plants; Cereal storage proteins; Grain protein composition; Mass spectrometry