Differentially expressed transposons between wild-type and <i>fpa-7</i>.

<p>(<i>A</i>) Differential expression of the transposable element gene (At5g10670) in <i>fpa-7</i>. (<i>B</i>) Read-through contiguous RNAs were validated by RT-PCR (red dashed line). Three biological replicates (1, 2 and 3) were used for each genotype: wild-type (WT) and <i>fpa</i>-7. UBIQUITIN <i>LIGASE 21</i> (<i>UBC21</i>) was used as a control. RT-PCR products were separated on agarose gels and stained with ethidium bromide. (<i>C</i>) Transcripts are either cleaved and polyadenylated in the annotated 3′UTR or at the intergenic sites, as determined by sequencing the cloned RT-PCR products. Red rectangles represent the 3′UTR specific to the read-through transcript and red lines represent 3′UTR introns. (<i>D</i>) Differential expression of the transposable element gene (At5g35935) in <i>fpa-7</i>. Recent re-annotation of At5g35935 <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1003867#pgen.1003867-Maher1" target="_blank">[12]</a>, <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1003867#pgen.1003867-Kannan1" target="_blank">[13]</a>, <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1003867#pgen.1003867-Garcia1" target="_blank">[28]</a>, <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1003867#pgen.1003867-Pontier1" target="_blank">[29]</a> defines two transcription units within it: the recently arisen pseudogene <i>psORF</i> and the transposon At5TE50260. DRS data reveal that silencing of <i>psORF</i> is lost in <i>fpa-7</i>. (<i>E</i>) RT-qPCR analysis of <i>psORF</i> in <i>fpa-7</i> and <i>fpa-8</i> mutant alleles. Silencing of <i>psORF</i> (p2 and p2b) is lost in <i>fpa-7</i> but not in <i>fpa-8</i>. Data are the means ± SEM obtained for three independent PCR amplifications of three biological replicates. The y-axis shows the fold change relative to WT (WT set to 1) after normalisation to <i>UBC21</i> gene expression. Location of the RT-qPCR amplicon is displayed on the left panel. <i>*</i>, <i>P</i><0.05; Student's t-test. Normalised reads mapping to the different loci are presented for WT and <i>fpa</i>. Genes are orientated 5′–3′; exons are denoted by rectangles, UTRs by adjoining narrower rectangles and introns by lines. Images of normalised read alignments were made using the Integrated Genome Browser <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1003867#pgen.1003867-Nicol1" target="_blank">[55]</a> and correspond to combined reads from the three sequenced biological replicates for each genotype.