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. 2018 Sep 10;9:1248. doi: 10.3389/fpls.2018.01248

Figure 3.

Figure 3

Quantification and subcellular distribution of hordeins at 12 and ≥20 DAP of endosperm development. (A,B) LFQ intensity of D-hordein and B3-hordein of aleurone 12 DAP (AL12) and 20 DAP (AL20), subaleurone 12 DAP (SA12), and ≥20 DAP (SA20) and starchy endosperm at 12 DAP (SE12) and ≥20 DAP (SE20). LFQ intensities of proteins were averaged over three replications. Bars represent standard deviation. For statistical analyses we performed a Student's t-Test (n = 3). The p-values indicate significant quantitative changes of hordeins. (C,D) Immunofluorescence of hordeins with anti-gliadin antibody shows morphological changes of the hordein-positive PBs (indicated by arrows) between 12 DAP (C) and ≥20 DAP (D) in subaleurone (SA) and starchy endosperm (SE). Note the very faint signal in aleurone (AL) at ≥20 DAP. The first two panels show overview pictures of the bright field and the fluorescence channel, respectively. Scale bar = 20 μm. Asterisks indicate starch granule. (E,F) Fluorescent signal quantification of anti-gliadin. Bar plots of the fluorescent intensity of anti-gliadin in all tissues at 12 and ≥20 DAP and in aleurone, subaleurone, and starchy endosperm separately at 12 and ≥20 DAP. Areas (n = 10–13 for 12 DAP; n = 7–11 for ≥20 DAP) were quantified for aleurone (AL), subaleurone (SA), and starchy endosperm (SE) from at least 2 slides with two sections. For statistical analyses we performed a Student's t-test. Bars represent standard deviation. Note the indicated p-values.