Functional Analysis of Chloroplast Iron Uptake and Homeostasis

Iron has a crucial role in plastid biology. Iron is a required cofactor for the operation of the photosynthetic functions and other metabolic pathways. Despite the importance of the iron homeostasis in chloroplasts, the functional analysis of the plastidial iron uptake and homeostasis still lack a consensus methodology. Here, we describe a sequence of subsequent techniques that can be applied in functional characterization of proteins involved in iron uptake and incorporation into chloroplasts as well as of the non-transport protein members of the chloroplast iron homeostasis. Since the ferrous iron ligation of bathophenantroline disulfonate is specific and not disrupted by the presence of other transition metals, it offers a simple way for iron quantification both in solubilized chloroplast samples as well as in ferric chelate reductase activity measurements.