One way to discover promoters is through an experiment called RNA polymerase protection assay, in which...

One way to discover promoters is through an experiment called RNA polymerase protection assay, in which RNA polymerase binds to a gene, is cross-linked to the DNA, and then the DNA is subjected to fragmentation, which breaks apart any DNA that is not protected by cross-linked protein. Explain why this would help to identify promoters in genes.

Expert Solution

Promoters are regions on DNA that initiate transcription of particular gene. Promoters may be located in an intergenic region. These regions can be immediately upstream of the gene of interest, or within or upstream of an adjacent open reading frame. The transcripts formed may be monocistronic or polycistronic. Promoter sequences determine which strand is transcribed.

RNA polymerase will bind to the Promoter with the other transcription factors. In eukaryotes, TATA box is present to which RNA polymerase bind. In prokaryotes, -10 and -35 sequences are present in the promoter region.

In RNA polymerase protection assay, the RNA polymerase will bind to the promoter region of the gene on the DNA. Cross linking will allow strong binding of RNA polymerase to the promoter. Fragmentation of DNA will remove any DNA that is not crosslinked as these are not protected or bound by the RNA polymerase. The unbound DNA is separated or purified as it is fragmented. Hence, the sample only will fragments that contain RNA polymerase bound to the any promoter. These fragments can be separated on a gel and then the sequence of the promoter can be identified by sequencing the fragments. The number of fragments will correspond to the number of promoter sites in the DNA.

gladiator answered 2 years ago

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