Skip to main content
. 2019 Oct;25(10):1298–1310. doi: 10.1261/rna.070425.119

FIGURE 1.

FIGURE 1.

Use of the AID system to conditionally deplete Spt5. (A) Western blot probed with anti-Flag and anti-Pgk1 as a loading control. Samples were taken before (T0) and 40 min (T40) after addition of auxin and β-estradiol. Spt5-AID* depletion was quantified and shown as the percentage mean of three biological replicates for T40 relative to T0 and normalized to the Pgk1 signal. Error bars, standard error of the mean. Gray crosses indicate the individual replicate values. (B) A diagram is drawn to scale, showing the positions of amplicons used for ChIP-qPCR analyses across each of the intron-containing genes ACT1, RPS13, ECM33. Exons are represented by gray rectangles and a scale bar of 1 kb is shown. (C) Anti-Flag ChIP followed by qPCR analysis of the intron-containing genes ACT1, RPS13, ECM33 without (−) auxin and β-estradiol (solid black line) or (+) 40 min after auxin and β-estradiol (dashed black line) addition to depleting Spt5-AID*-6Flag. The x-axis of each graph shows the amplicons used for ChIP-qPCR analysis. The data are presented as the mean percentage of input relative to the first amplicon of each gene for at least three biological replicates. Error bars, standard error of the mean. Asterisks show the statistical significance (Student's unpaired t-test). (*) P < 0.05, (**) P < 0.01, and (***) P < 0.001. Not significant, P > 0.05. Gray crosses indicate the individual replicate values without auxin and β-estradiol, and gray circles indicate the individual replicate values 40 min after auxin and β-estradiol addition.