RNA interference-mediated knock-down of α1,3-fucose and β1,2-xylose in rice N-glycans
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RNA interference (RNAi) was performed for the down-regulated expression of two endogeneous core α1,3-fucosyltransferase (FucT) and β1,2-xylosyltransferase (XylT) genes in rice (Oryza sativa). We constructed three RNAi vectors for the suppression of two glycosyltransferase transcriptions and transformed them into the rice plants by Agrobacterium. The transgenic plants were selected by PCR and down-regulated expressions of FucT and XylT in the mRNA level were verified by RT-PCR and qRT-PCR. In addition, structural properties of N-glycans from glycoproteins in transgenic rice leaves were analyzed by MALDI-TOF MS and were compared with those of wild-type rice leaves. The compositional change of plant-specific N-glycans in total endogenous rice leaf glycoproteins was induced as consequence of the knock-down of two N-glycan maturation glycosyltransferases by using RNAi. Compositional change of N-glycans in three selected RNAi suppression plants was observed to decrease the ratio of three major hybrid, hybrid bisected and paucimanosidic-type N-glycans: GlcNAc3Man3Xyl1Fuc1, GlcNAc4Man3Xyl1-Fuc1 and GlcNAc2Man3Xyl1Fuc1.
KeywordsRice N-glycan β1,2-xylosyltransferase α1,3-fucosyltransferase RNA interference
Polymerase chain reaction
Quantitative real-time PCR
Matrix assisted laser desorption ionization-time of flight mass spectrometry
National center for biotechnology information
Basic local alignment search tool
This project was supported by grant (to B.S.H.) from National Academy of Agricultural Science (PJ008541). This study was supported by 2014 Postdoctoral Fellowship Program (to H.Y.C.) of National Academy of Agricultural Science, RDA, Korea.
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