Date of Award
Program or Major
Doctor of Philosophy
Subhash C Minocha
Ornithine decarboxylase (ODC), arginine decarboxylase (ADC), and S-adenosylmethionine decarboxylase (SAMDC) are three of the key regulatory enzymes involved in the biosynthesis of polyamines (putrescine, spermidine and spermine). To modulate the biosynthesis of putrescine, carrot (Daucus carota L.) cells were transformed with Agrobacterium tumefaciens strains containing 3$\sp\prime$-truncated mouse ornithine decarboxylase (ODC) cDNA under the control of a CaMV 35S promoter. Neomycin phosphotransferase gene linked with nopaline synthase promoter was used to select transformed cell lines on kanamycin. While the non-transformed cells contained no ODC, high levels of ODC activity were observed in the transformed cells. A significant increase in the cellular levels of putrescine in transgenic cells as compared to control cells was observed. Spermidine levels, however, remained unaffected. Not only did the transformed cells exhibit improved somatic embryogenesis in the auxin-free medium, they also regenerated embryos in the presence of inhibitory levels of 2,4-D. These cells acquired tolerance to $\alpha$-difluoromethylarginine (a potent inhibitor of arginine decarboxylase) at concentrations that inhibit growth as well as embryogenesis in non-transformed carrot cells.
Transformation of carrot cells with a human SAMDC cDNA lead to increased production of SAMDC enzyme. This increase in the biosynthesis of SAMDC translated to an increase in the cellular levels of spermidine and a decrease of putrescine. The transgenic cells were highly embryogenic and also tolerant to low levels of methylglyoxal bis(guanylhydrazone).
Bastola, Dhundy Raj, "Genetic engineering of the polyamine biosynthetic pathway and somatic embryogenesis in carrot (Daucus carota L)" (1994). Doctoral Dissertations. 1812.