diagram showing NOS-bar (B) and rbcS-35S (C) regions for methylation analysis on T-DNA of the unmodified 35S-sGFP construct. (B, C) Cytosine methylation status of NOS promoter (black) with bar coding (blue) (B) and Arabidopsis rbcS terminator (green) with 35Spro (black) (C) regions. The percentage of methylated cytosines is represented by bar charts (red, CpG; green, CpWpG; black, CpHpH), and the position of each cytosine is represented below. Positions of the start codon and TATA-box are indicated in pink and aqua, respectively, and positions of the known elements within the promoter regions are indicated by different colors. Found at: doi:10.1371/journal.pone.0009670.s004 (0.13 MB PDF) Figure S5 CpHpH methylation states of 35S(Dcore) lines in a time-series of culture. Percent methylation at CpHpH sites of the 35S(Dcore) lines, from which the genomic DNAs were obtained on Oct. 16, 2006 (black bars) and Mar. 22, 2007 (gray bars; the same data is represented on Figure 4), respectively, is shown. Analyzed regions of the 35S(Dcore) promoter with sGFP (black bar; indicated by arrows) are represented below. Positions of cytosines corre- sponding to the sequence from 2148 to 285 of the unmodified 35S pro are indicated by red and pale red, respectively. Found at: doi:10.1371/journal.pone.0009670.s005 (0.11 MB PDF) Figure S6 Small RNA and genomic DNA analyses to identify the endogenous 35S enhancer sequence in gentian. (A) Small- molecule RNAs from leaf tissues of wild-type (WT) and unmodified 35S-sGFP line #8 of gentian plants, and 35S-sGFP tobacco line #26 (C) were electrophoresed and hybridized with DIG-labeled 35S enhancer (upper) and 5S rRNA (lower) probes. Ethidium-stained gel bands serve as loading controls. 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Mol Gen Genet 244: 219–229. PLoS ONE | www.plosone.org 10 March 2010 | Volume 5 | Issue 3 | e9670