Localization of Vitamin E Constituents in Transgenic Cotton (Gossypium hirustum L.) Embryos

Cottonseed oil is a naturally rich source of vitamin E (tocopherols), with alpha- and gamma- tocopherols comprising nearly all of the tocopherols in cottonseed oil. Most dicot seeds like cotton lack the tocotrienols (the unsaturated versions of the tocopherols) that are found in monocot seeds. Transgenic cotton plants with a gene construct, Napin::HvHGGT (containing the Hordeum vulgare (Hv) homogentisate geranylgeranyl transferase (HGGT) gene/coding sequence under the control of Brassica napus Napin, seed-specific promoter) were produced by Agrobacterium-mediated transformation of embryogenic cell lines of G. hirsutum variety Coker 312. Biochemical analysis of transgenic cottonseeds showed about 3-fold increase in the accumulation of total vitamin E (tocopherols + tocotrienols) which included a substantial proportion of tocotrienols in comparison to non-transgenic Coker 312. Tocotrienols are completely absent in non-transgenic cotton Coker 312. Matrix assisted laser desorption ionization- mass spectrometry (MALDI-MS) imaging and localization of vitamin E constituents in cotton embryo tissue sections of transgenic and non-transgenic cotton embryos were carried out. As expected, MALDI-MS image analysis of Coker 312 embryos showed presence of α-tocopherol restricted to the cotyledonary tissue while γ-tocopherol was found in the axial tissue. Transgenic embryos showed the same distribution in tocopherols as Coker 312, but strong signals for the presence of γ-tocotrienol throughout the embryo, thus revealing heterogeneous distribution of α- and γ-tocopherol, and confirming the accumulation of γ-tocotrienol in transgenic cotton embryos.