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. 2017 Mar 14;6:e23001. doi: 10.7554/eLife.23001

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© 2017, Pateraki et al

This article is distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use and redistribution provided that the original author and source are credited.

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Figure 5. — Total ion chromatograms (TIC) of extracts expressing the 13R-manoyl oxide biosynthesis genes (CfCXS, CfGGPPS, CfTPS2, CfTPS3), in combination with (from the top) water (-), CfCYP76AH8 + CfCYP76AH11 + CfCYP76AH16, CfCYP76AH17 + CfCYP76AH11 + CfCYP76AH16, and CfCYP76AH15 + CfCYP76AH11 + CfCYP76AH16 are shown. Hydroxylated 13R-manoyl oxide-derived diterpenoids (marked with grey bars) and their identity including their molecular formulas were confirmed by accurate mass (5 ppm tolerance, Supplementary file 1). Compounds present in trace amounts are not marked. The identity of 1,11-dihydroxy-13R-manoyl oxide (5d), 9-deoxydeacetylforskolin (10b) and 1,9-dideoxydeacetylforskolin (7h) was confirmed by NMR analysis (Figure 4 and Tables 1 and 2), whereas the identity of deacetylforskolin (13b) was confirmed by comparison to an authentic chemically synthesized standard. No 13R-manoyl oxide-derived diterpenoids were identified in the water control (-). For each combinaton, extracts from leaves of three different N. benthamiana plants have been analyzed and representative chromatograms are shown.

DOI: http://dx.doi.org/10.7554/eLife.23001.015

Figure 5—figure supplement 1. — Total ion chromatograms (TIC) of extracts expressing the 13R-manoyl oxide biosynthesis genes (CfCXS, CfGGPPS, CfTPS2, CfTPS3), in combination with (from the top) water (-), CfCYP76AH8 + CfCYP76AH11 + CfCYP76AH16, CfCYP76AH17 + CfCYP76AH11 + CfCYP76AH16, and CfCYP76AH15 + CfCYP76AH11 + CfCYP76AH16 are shown. Hydroxylated 13R-manoyl oxide-derived diterpenoids (marked with grey bars) and their identity including their molecular formulas were confirmed by accurate mass (5 ppm tolerance, Supplementary file 1). Compounds present in trace amounts are not marked. The identity of 1,11-dihydroxy-13R-manoyl oxide (5d), 9-deoxydeacetylforskolin (10b) and 1,9-dideoxydeacetylforskolin (7h) was confirmed by NMR analysis (Figure 4 and Tables 1 and 2), whereas the identity of deacetylforskolin (13b) was confirmed by comparison to an authentic chemically synthesized standard. No 13R-manoyl oxide-derived diterpenoids were identified in the water control (-). For each combinaton, extracts from leaves of three different N. benthamiana plants have been analyzed and representative chromatograms are shown.

DOI: http://dx.doi.org/10.7554/eLife.23001.015