Institut for Agroøkologi

Titel på PhD projekt:
Iron biofortification of the wheat endosperm

Micronutrient deficiency is of global concern. The WHO estimates the number of anaemic people worldwide to be more than two billion, primarily women and children and that approximately 50% of all anaemia can be attributed to iron deficiency, this call for solutions.

Anaemia is an indicator of undiversified and poor nutrition and is prevalent among poor populations in undeveloped or developing countries. In these countries the poverty and lack of infrastructure in the rural areas consequently obstructs and makes a food or vitamin based supplementation inefficient. Another solution, breeding of stable crops for the trait of iron content, has shown to be difficult, due to low diversity for this trait among the modern cultivars. In contrast research using molecular approaches show promising results. Expression of the iron sequestering protein ferritin in the endosperm of wheat and rice raise the iron content in the grain by a factor 2-3. It is expected that the extra iron storage capacity caused by ferritin will forces the transfer of additional amounts of iron from the aleurone layer into the endosperm.

Wheat genome has two ferritin genes each with three homoeoalleles. My aim is to produce transgenic wheat lines that overexpress one of the wheat’s own ferritin proteins in the grain endosperm using the high molecular weight glutenin promoter 1Dx5. We have the ambition to produce high iron transgenic lines that can be used directly in breeding. Therefore we want to transform modern wheat cultivars and produce transformants that are free of the selection marker gene and vector backbone DNA that are impeding the legislation. Gene gun is the appliance we will use to co-transform the cassettes of the gene of interest and marker gene. Afterwards offspring segregation will help us to screen and select transformants that have only the gene of interest into their genome. This is possible due to halving of chromosome pairs in gametes. In case of linkage between inserted cassettes, segregation is made possible via crossing-over.

The protein storage vacuoles in the aleurone layer are major storage sites for iron in the grain. Here the iron is stored together with phytate, a phosphate storage compound with significant anti-nutritional properties. Complexed to phytate iron has a low bioavailability that impedes its uptake. To try to overcome the problem we will cross the high ferritin transgenic lines with lines expressing phytase to improve the bioavailability of iron in the digestive tract of humans and livestock.

The other part of my project is transcriptome analysis on grains from wheat plants under different iron treatments. The Laser Capture Microdissection procedure will be used for sampling from cryosectioned grains. The transcription profile of iron transporters in different grain parts of the grain will be used to predict a model for iron translocation into the grain during grainfilling and will be useful to identify new candidate genes involved in the iron pathway into the grain.

The final part of my project will be subjected the modulation of key iron transporters to redirect iron deposition from the aleurone storage vacuole to the endosperm. The Vacuolar Iron Tranporter (Vit1) and the Nramp transporter is respectively hypothised to pump iron into and out of the protein storage vacuole. RNAi will be used to downregulate the genes and grain specific promoters will be used to produce overexpressores. In this concept, cloning of wheat specific promoters, e.g. LTP2 promoter, the Vacuolar Iron Tranporter (Vit1) and Nramp genes and the modulation of these iron transporters are the main targets. It is expected that the downregulation of Vit gene(s) and overexpression of Nramp genes, could be as a potential stimulus to leak iron out of aleurone and into the endosperm.