Molecular dissection of species-specific transgene silencing

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Molecular mechanism of the 35S promoter-specific DNA methylation in gentian was investigated in this study. Two distinct de novo methylated regions were identified after the methylation analysis of an entire T-DNA segment in transgenic gentian. Both of the regions contain two consensus sequences, to which gentian nuclear extracts could bind, and yeast one-hybrid screening identified candidates for binding to the sequences. Additionally, relationship between DNA methylation and histone modification was investigated using transgenic gentian cell suspension culture, and the results showed that de novo methylation of the 35S region occurred in connection with histone modification. To investigate whether the 35S promoter-specific DNA methylation exists in other plant species, transgenic lettuce plants having 35S promoter were produced. As the result, suppression of the transgene expression and hypermethylation of the 35S promoter region were found in the transgenic lettuce plants.