Chlamydomonas reinhardtii Fea1 Protein Facilitates Iron Uptake in Transgenic Plants
Creators:Chiu, Wai Ting
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Publisher:The Ohio State University
Series/Report no.:The Ohio State University. Department of Microbiology Honors Theses; 2007
Anaemia caused by iron deficiency is a significant public health problem in numerous developing countries, affecting about one third of the world’s population. Iron deficiency often leads to low productivity, compromised immunity, reduced cognitive development in children and increased mortality during pregnancy. These health conditions result largely from decreased hemoglobin and myoglobin levels in the blood. Since the most abundant source of iron available for human consumption resides in dietary plants, improving iron content in the diet may be one of the most effective methods to eliminate iron deficiency anaemia. To increase iron assimilation in plants, our group proposed transformation of Arabidopsis thaliana (ecotype Columbia) with the Chlamydomonas fea1 gene using two different plasmids, where its expression is under the regulation of the constitutive 2X 35S CAMV or the root-specific patatin promoter. The fea1 gene of Chlamydomonas fea1 encodes an iron transport protein that is induced in Chlamydomonas under high carbon dioxide, iron depletion and cadmium stress conditions. Fea1 transformants were confirmed by PCR and RT-PCR. Seedling emergence studies showed that the transgenic plants carrying the FEA1 construct expressed under the control of either promoter performed better than wild-type plants under iron limiting conditions. Moreover, root pH indicator dye experiments revealed a reduced proton extrusion under iron stress compared with WT, indicative of an increased ability to uptake iron. To examine further the role of the fea1 gene product in iron transport, the Arabidopsis thaliana IRT1 mutant, having impaired iron assimilation, was transformed with the Chlamydomonas fea1 gene. The results revealed that the fea1 gene complemented the IRT1 mutant by imparting the transgenic plants with the capability to survive in soil without exogenous iron supplementation, which is required for the Arabidopsis thaliana IRT1 mutant. In addition, chlorophyll fluorescence kinetic studies conducted with low iron grown wild-type and transgenic plants showed a lower fluorescence yield and normal fluorescence decay kinetics in plants expressing the fea1 gene relative to non-transgenic plants. These results are indicative of higher photochemical efficiency as well as normal non-photochemical quenching activity, indicative of a functional cytochrome b6f complex. Further studies to be conducted include: tissue-specific RT-PCR analysis of transformants, and analysis of the iron content of transgenic plants grown under iron stress and sufficient conditions.
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