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Fig. 2 | Plant Methods

Fig. 2

From: A high-throughput screening method to identify proteins involved in unfolded protein response of the endoplasmic reticulum in plants

Fig. 2

Reporter choice and optimization of unfolded protein response (UPR) induction. a Four reporter constructs were tested for enhanced yellow fluorescent protein (eYFP) upregulation after UPR induction by 5 μg/mL tunicamycin (Tm) infiltration. Subsequent tests were carried out using the pBIP1::eYFP construct. b Sampling at 6, 12, 24, and 48 h after Tm infiltration was tested. In the 48 h samples, the gain value of the fluorescence detector was lowered from 100 to 90. The 24 h timepoint was used for further tests. c Different Tm concentrations were tested for UPR induction. All future tests were done with 5 μg/mL Tm. d Three optical density at 600 nm (OD600 nm) ratios of A. tumefaciens strains were tested for optimal eYFP induction after Tm infiltration, and e candidate protein expression levels, using mCherry (mCh) as a reference. In both d and e, the A. tumefaciens strain carrying the pBIP1::eYFP reporter construct was co-infiltrated with an A. tumefaciens strain carrying a p35S::mCh-P2A-mCh control construct. Error bars represent standard deviation, asterisks represent statistically significant differences (one-way ANOVA or t-test) between samples: *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001, and n.s. not significant. Lower case letters represent differences between treatments among samples infiltrated with the same A. tumefaciens suspension, while capital letters represent differences within the same treatment among samples infiltrated with different A. tumefaciens suspensions in a two-way ANOVA test (P ≤ 0.05). a.u. arbitrary units

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