Increasing seed oil production is a major goal for global agriculture to meet the strong demand for oil consumption by humans and for biodiesel production. Previous studies to increase oil synthesis in plants have focused mainly on manipulation of oil pathway genes. As an alternative to single-enzyme approaches, transcription factors provide an attractive solution for altering complex traits, with the caveat that transcription factors may face the challenge of undesirable pleiotropic effects. Here, we report that overexpression of maize (Zea mays) LEAFY COTYLEDON1 (ZmLEC1) increases seed oil by as much as 48% but reduces seed germination and leaf growth in maize. To uncouple oil increase from the undesirable agronomic traits, we identified a LEC1 downstream transcription factor, maize WRINKLED1 (ZmWRI1). Overexpression of ZmWRI1 results in an oil increase similar to overexpression of ZmLEC1 without affecting germination, seedling growth, or grain yield. These results emphasize the importance of field testing for developing a commercial high-oil product and highlight ZmWRI1 as a promising target for increasing oil production in crops.

Plant oil is an important renewable resource for biodiesel production and for dietary consumption by humans and livestock. Through genetic mapping of the oil trait in plants, studies have reported multiple quantitative trait loci (QTLs) with small effects, but the molecular basis of oil QTLs remains largely unknown 1 , 2 , 3 , 4 , 5 . Here we show that a high-oil QTL ( qHO6 ) affecting maize seed oil and oleic-acid contents encodes an acyl-CoA:diacylglycerol acyltransferase (DGAT1-2), which catalyzes the final step of oil synthesis. We further show that a phenylalanine insertion in DGAT1-2 at position 469 (F469) is responsible for the increased oil and oleic-acid contents. The DGAT1-2 allele with F469 is ancestral, whereas the allele without F469 is a more recent mutant selected by domestication or breeding. Ectopic expression of the high-oil DGAT1-2 allele increases oil and oleic-acid contents by up to 41% and 107%, respectively. This work provides insights into the molecular basis of natural variation of oil and oleic-acid contents in plants and highlights DGAT as a promising target for increasing oil and oleic-acid contents in other crops.

Glucosyltransferases (GTFs, EC.2.4.1.5) are bacterial enzymes that catalyze the polymerization of glucose residues from sucrose, leading to the production of high molecular weight glucan with α-1,3 /α-1,6 linkages. Such glucans, with many potential food and industrial applications, do not normally exist in higher plants. We fused a mutant form of the gtfD gene from Sreptococcus mutans with the maize (Zea mays L.) chloroplastic Brittle 1 transit peptide for amyloplast targeting. This construct, driven by the ubiquitin promoter, was introduced into maize by Agrobacterium-mediated transformation. We developed a novel HPLC-based method that enabled us differentially to distinguish transgene glucan from other endogenous polysaccharides in maize kernels. Using this method, we screened over 100 transgenic plants for the presence of GTF-produced glucan whose content varied between 0.8 and 14% of dry weight in the mature transgenic seeds. The mature transgenic plants were indistinguishable from wildtype plants in growth rate and morphology. Furthermore, starch granule size in the transgenic maize kernel was unaffected by the accumulation of the foreign polysaccharide. Mutation in Sh2, which encodes a subunit of ADP-glucose pyrophosphorylase, had no effect on glucan accumulation caused by gtfD expression. Our results indicated that high levels of novel carbohydrate polymer can be accumulated in crop plants through transgene technology.

Phytic acid in cereal grains and oilseeds is poorly digested by monogastric animals and negatively affects animal nutrition and the environment. However, breeding programs involving mutants with less phytic acid and more inorganic phosphate (P(i)) have been frustrated by undesirable agronomic characteristics associated with the phytic acid-reducing mutations. We show that maize lpa1 mutants are defective in a multidrug resistance-associated protein (MRP) ATP-binding cassette (ABC) transporter that is expressed most highly in embryos, but also in immature endosperm, germinating seed and vegetative tissues. Silencing expression of this transporter in an embryo-specific manner produced low-phytic-acid, high-Pi transgenic maize seeds that germinate normally and do not show any significant reduction in seed dry weight. This dominant transgenic approach obviates the need for incorporating recessive lpa1 mutations to create maize hybrids with reduced phytic acid. Suppressing the homologous soybean MRP gene also generated low-phytic-acid seed, suggesting that the strategy might be feasible for many crops.

This chapter describes a stepwise protocol to achieve success in genetic transformation of maize using Agrobacterium tumefaciens as a DNA delivery system. Researchers will be able to effectively transform immature embryos of Hi-II and related genotypes with this protocol. The outcome of the transformation process will be transgenic embryogenic callus tissue, transgenic plants, and transgenic progeny seeds. Recommendations for molecular confirmation and evaluation of transgenic tissue/plants are also provided.

The maize (Zea mays L.) endosperm consists of an epidermal like layer of isodiametric aleurone cells surrounding a central body of starchy endosperm cells. In disorgal1 (dil1) and disorgal2 (dil2) mutants the control of the mitotic division plane is relaxed or missing, resulting in mature grains with disorganized aleurone layers. In addition to orientation of the division plane, both the shape and size of the aleurone cells are affected, and often more than one layer of aleurone cells is present. Homozygous dil1 and dil2 grains are shrunken due to reduced accumulation of starchy endosperm and premature developmental arrest of the embryo, and mature mutant grains germinate at a very low rate and fail to develop into plants. However, homozygous mutant plants can be obtained through embryo rescue, revealing that both mutants have an irregular leaf epidermis as well as roots with a strongly reduced number of root hairs and aberrant root hair morphology. Our results suggest the presence of common regulatory mechanisms for the control of cell division orientation in the aleurone and plant epidermis.