Abstract
In this investigation we analyzed the effect of DNA methylation on gene expression in tobacco protoplasts. Methylation of the chimeric GUS gene (CaMV 35s/GUS/NOS) with the HpaII methyltransferase had no effect on transient and on stable GUS gene expression. The chimeric gene was then cloned as single-stranded DNA. In vitro second-strand DNA synthesis was performed with 5-methylcytosine, leading to methylation of every cytosine in one DNA strand. After transfection this DNA was totally inactive. Control experiments revealed that the single-stranded DNA was as active as the double-stranded form.
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