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Fig. 3 | BMC Plant Biology

Fig. 3

From: The organ-specific differential roles of rice DXS and DXR, the first two enzymes of the MEP pathway, in carotenoid metabolism in Oryza sativa leaves and seeds

Fig. 3

Schematic representation of the carotenoid pathway, the binary vectors used in this study, and transgene expression in leaves and seeds of rice plants. a Built-in pathway for carotenoid biosynthesis in rice plants; CRTI, Pantoea annatis desaturase; DMAPP, dimethylallyl diphosphate; DXP, deoxyxylulose 5-phosphate; DXR, DXP reductoisomerase; DXS, DXP synthase; IPP, isopentenyl diphosphate; GGPP, geranylgeranyl pyrophosphate; G3P, glyceraldehyde 3-phosphate; LCYB, lycopene β-cyclase; MEP, methylerythritol 4-phosphate; OsDXR, rice DXP reductoisomerase; OsDXS2, rice DXP synthase; PSY, phytoene synthase; stCrtI, rice codon-optimized synthetic gene encoding the Pantoea CrtI gene; stPsy, rice codon-optimized synthetic gene of the Capsicum gene encoding PSY; stPAC, a recombinant gene of stPsy and stCrt linked with 2A, which is the rice codon-optimized foot-and-mouth disease virus 2A peptide. b Diagrams of the four vectors for rice transformation of OsDXS2 and OsDXR with the pGlb::stPAC vector that was previously used to generate a carotenoid-intensifying trait in rice endosperm [32]. Bacterial attachment attB sites needed for Gateway cloning are marked with hatched boxes. BR, right border; BL, left border; PGD1, rice phosphogluconate dehydrogenase promoter; OsDXS2, rice DXP synthase 2 gene; OsDXR, rice DXP reductoisomerase gene; PinII, 3′ region of the potato proteinase inhibitor II gene; BAR, Bialaphos-resistant gene cassette; Glb, rice globulin promoter; stPAC, a recombinant gene of stPsy-2A-stCrt. The primer locations used in vector construction, PCR, and qRT-PCR for the transgene analyses in Additional file 3: Figure S3a, Fig. 3c, and Fig. 3d are indicated as arrows, and their information is listed in Additional file 4: Table S7. c Transgene expression levels of OsDXS2 and OsDXR were examined by qRT-PCR using total RNA isolated from 10-day-old leaves. d Transgene expression levels of OsDXS2, OsDXR, and stPAC were examined by qRT-PCR using total RNA purified from unpolished mature seeds 40 DAF. All results using gene-specific primer pairs F7/R7 for OsDXS2, F8/F7 for OsDXR, and F9/R9 for stPAC were calculated as the mean of three replicates and normalized to the expression of the OsUbi5 gene, which was amplified using a U5F/U5R primer pair. The primers are indicated in Fig. 3b and listed in Additional file 4: Table S7. NT is a non-transgenic wild type of Oryza sativa L. cv. Ilmi, different varieties of transgenic plants are represented in different colors, such as blue, light-blue, orange, light-orange and yellow in the bar graph, and the X-axis labeling consists of three independent-transgenic plant lines

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