Why introduction of an extra copy of a gene related to pigmentation causes RNA interference in Petunia?

Question

RNAi became famous after Fire and Mello experiment in C.elegans; however, it had been observed before. In the 80's, Jorgensen was trying to increase pigmentation of Petunia flowers by introducing extra copies of chalcone synthetase gene, which encodes an enzyme involved in anthocyanin synthesis.

However, this experiment lead to post transcriptional gene silencing (PTGS) by a mechanism that was not understood at that time, called RNA interference.

Here you might find more information about the experiment (including the vector used to introduce the gene).

I know how RNAi works by the classical pathway; however, I didn't understand how an extra copy of the gene could generate gene silencing. Introducing of DNA sequences has been used to knock downs target genes, however, generally these sequences are planned so that they will form a hairpin which is recognised by Dicer; probably, the researcher did not use such a sequence. Moreover, exogenenic genes are widely used without resulting in unplanned RNAi.

Does someone know more information about what possibly happened in the experiment to produce RNAi?

Answer 1

The answer is in the slides you provided a link to. The key fact is that Craig & Andy did careful controls to show that it was the presence of dsRNA in both the sense control, and in the anti-sense experimental sample that was responsible for the RNAi-mediated knock-down. Actually, Ken Kemphues showed this earlier in a Nature paper (the control also knocked down gene expression) but did not pursue the mechanism.

Therefore, I infer that the transgenic gene expression construct introduced into the petunias must also give rise to dsRNA. I am less familiar with the consequences of introducing foreign DNA into a plant cell (e.g., does a single copy become integrated, multiple copies?, is it maintained extrachromosomally?)