Metal content analysis
For total Fe content analysis, plant samples were collected and washed
with 2 mM CaCl2. The samples were firstly dried at 105℃
for 0.5 h and subsequently oven-dried at 70℃. The oven-dried samples
were ground into powder and its dry weight was recorded. Next, the
powder was put into 50 mL conical flask dispersed in 5 mL concentrated
H2SO4, incubated at room temperature
(~24℃) for 12 h. Finally, the samples were heated to
330℃ and digested by slowly adding 10 mL 30%
H2O2 until it was clear (Smolders et al.
1997). The digested liquid was filtered with 0.45 µM filter and filtrate
was diluted with ddH2O. Total Fe content was assessment
by inductively coupled plasma optical emission spectrometer (ICP-OES,
iCAP 7400, Thermo Fisher Scientific, Bremen, Germany).
Water-soluble Fe was extracted according to Cassin et al . (2009).
Briefly, samples were weighed and ground in liquid nitrogen. Then, 10 mL
deionized water was added at room temperature. Subsequently, it was
centrifugation for 10 min and the supernatant was collected. The Fe
concentration of the supernatant was measured using the same method as
total Fe.