a. Top: Diagram of SUVH2 with Zn Finger (ZF) inserted immediately before the HA tag. Bottom: Schematic of FWA gene showing the two small and two large repeats (blue arrows), the regions amplified by PCR (promoter and transcript: green lines), the start and direction of transcription (red arrow), and the sites bound by the ZF (indicated by two orange arrows).
b. Flag-ChIP in WT versus ZF-KYP (flag-tagged) showing enrichment at FWA in both the promoter and transcript region (see above).
c. Percent methylation at each C in the FWA repeat region from three individual T1 plants. Percent methylation was determined from 20-25 clones of bisulfite-treated DNA.
d. BS-Seq of FWA from a Basta-resistant line containing the ZF-SUVH2 transgene and two Basta-sensitivie siblings which had lost the ZF-SUVH2 transgene.
e. Pull-down of DRD1-Flag with ZF-SUVH2. Input: DRD1-Flag extract from Arabidopsis; Beads-mock: elution from DRD1-Flag pull-down using HA-magnetic beads pre-bound with Nicotiana benthamiana extract; Beads-ZF-SUVH2: elution from DRD1-Flag pull-down using HA-magnetic beads pre-bound with Nicotiana benthamiana ZF-SUVH2 extract. Top panel: Flag blot; bottom panel: HA blot.