T. E. Finn et al. 5 SI

FIGURE S4.—Bisulphite PCR analysis of a chloroplast gene encoding psaA protein using the same bisulphite-treated

genomic DNA as for the analysis of 35S promoter methylation in Figure 4A and C. (A). The sequence of the analysed region of the chloroplast DNA (157 bp; reverse complement of nt. 880-1036 of ATCG00350). Letters in red indicate the sequences

against which the forward and reverse bisulphite PCR primers are designed. Pink letters indicate the region of which the

bisulphite sequencing trace files are shown below in C. Underlined letters correspond to the two Msel sites created when the

cytosines are converted to thymines. (B). Msel restriction sites in bisulphite converted DNA. (C). Sequencing trace files of the

region spanning the two Msel sites in bisulphite-treated chloroplast DNA from the T2 lines showing significant 35S promoter

methylation. Note that the level of cytosines (blue peaks) is minimal in comparison with that of thymines, indicating efficient

and uniform bisulphite conversion.