Abstract
We introduced the inverted repeat (IR) silencer DNA construct to trigger RNAi in a unicellular green alga Chlamydomonas reinhardtii, which is targeted aadA mRNA Robust gene silencing was observed at some transformants immediately after transformation, however when the efficiency was reexamined after long period of continuous culture and subsequent single colony isolation, RNAi transformants showed heterogeneously decreased efficiency of RNAi. Prominent negative correlation between the amounts of accumulated aadA hairpin RNA and the RNAi efficiency was demonstrated by Northern blot analysis. Methylated cytosines were heavily accumulated in the IR region of silencer DNA and the modified cytosine ratio negatively correlated with accumulated hairpin RNA. Moreover, treatment of histone deacetylase inhibitor, immediately increased the accumulated hairpin RNA. Taking together, we propose that IR DNA structure of silencer construct induced CG-methylation at the region, then histone modification change followed to repress the transcription of the hairpin RNA.
