Recruitment of RNA-Directed DNA Methylation to Transformed Direct Repeats

<div><p>(A) Silencing of endogenous <i>FWA</i> by transformed <i>FWA</i> requires tandem repeats in the incoming transgene. Wild-type (WT) L<i>er</i> or <i>fwa-1</i> plants were transformed with a complete <i>FWA</i> transgene or the deletion variants shown at right. <i>FWA</i> contains two tandem repeats, which are depicted as two sets of arrows. Parental <i>fwa-1</i> plants are uniformly late flowering. Early flowering in <i>fwa-1</i> transformants occurs when endogenous <i>fwa</i> is silenced by de novo DNA methylation. Plants were scored as early- (17 leaves or fewer) or late-flowering (18 leaves or more) by comparison to untransformed L<i>er</i> and <i>fwa-1</i>.</p> <p>(B) <i>FWA</i> transgenes lacking tandem repeats do not cause late flowering when transformed into <i>rdr2–1</i>. The full <i>FWA</i> gene, “<i>FWA</i> repeats deleted” transgene, and “<i>FWA</i> single copy” transgenes are diagrammed. Wild-type Col and <i>rdr2–1</i> plants were transformed with each transgene, and flowering time was assayed in the T1 generation.</p> <p>(C) De novo DNA methylation of transformed <i>FWA</i> requires tandem repeat character. Wild-type Col plants were transformed with full-length <i>FWA</i> or with single-copy <i>FWA</i>. DNA methylation of the <i>FWA</i> transgene was assayed by bisulfite sequencing.</p>