You would like to transcribe a eukaryotic gene in vitro (ie. in a test tube in t

Question

You would like to transcribe a eukaryotic gene in vitro (ie. in a test tube in the lab). Your goal is to achieve a high level of transcription. To your test tube, you add the DNA template for this gene, which contains the following: a TATA box sequence located –25 to –30 bp upstream of the transcription start site, the sequence for the gene, and terminator sequences. You also add RNA polymerase and the transcription factors that make up the basal transcriptional machinery. However, after doing your experiment, you do not get very high levels of transcription. What do you think might be the cause of this and what corrective measures would you take to increase the level of transcription?

Explanation / Answer

The reason for not getting the high levels of transcription as the nucleotides and ions were not added in the solution.

To rectify the mistake you should add NTPs and Mg+2 ions to get high level of transcription.

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