We discussed several specific cis-elements in mRNAs that regulate splicing, stability, decay, localization, and translation. However, it is likely that many other uncharacterized cis-elements exist. One way in which they may be characterized is through the use of a reporter gene such as the gene encoding the green fluorescent protein (GFP) from jellyfish. GFP emits green fluorescence when excited by blue light. Explain how one might be able to devise an assay to test for the effect of various cis-elements on posttranscriptional gene regulation using cells that transcribe a GFP mRNA with genetically inserted cis-elements.

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Hi, everyone. Welcome back. Here's our next question. Which of the following techniques is used to study the effects of cis elements on post transcription, all gene regulation. So let's recall what CIS elements are. So, cis elements are non coding DNA sequences that regulate the expression of genes on the same chromosome that they are located on. So you have these non coding DNA sequences and the cis part means they're on the same chromosome and they're involved in post transcription and gene regulation of those genes. So if we want to study the effects of the cis elements, we need a little more information than just um what level the M RNA is expressed at because there's a lot of different factors and you carry outs that go into post transcription and gene regulation. So if you want to tease out specifically the effect of just these cis elements, um you need a slightly different technique. So let's look at our answer choices um noting that we do have to S D all of the above and when we look at them, um Choice A is RNA sequencing and Choice B is northern body. Both of these are not going to be our answer because they both involve just looking at the M R N A expression. So in in RNA sequencing, you'd be looking at the M RNA in your sample and sequencing to see which RNA sequences have been expressed. Then northern blotting is a technique where you separate RNA fragments on a gel. And then you use labeled C DNA probes complementary to the RNA sequences of interest to identify where those RNA fragments are. So you wash your gel with these labeled C DNA fragments and they will bind to and hybridize with any RNA fragments that are complementary to theirs sequence. So that allows you to look for which are, if there's a specific gene being expressed in your sample, you can find that, but that's not going to tell us that will tell us the final level of RNA expression and which are are have been transcribed, but it won't tell us the effect of a specific CIS element. So in that case, you need something a little more nuanced, which it will just look at that element in isolation. So for that, we're going to need choice D and we can also eliminate choice D, all of the above which since we have no, the ambi are incorrect. So to look at these elements, we need to look at choice, see reporter gene assay. And this involves the use of a reporter gene as its name says, that will allow you to look at the effect of this, this element on the reporter gene. So you use something like um a, a gene like G F P green fluorescent protein. Um You take the cis elements from your gene of interest, you have them on the D N A that you're going to create and transfer it into a cell. And so the G F P will be under the control of the cis elements. You're trying to study, then you transfer effect that plasma that you've created with the CIS element and the G F P gene into a cell incubated with the appropriate re agents. And then you look at how the result inflorescence differs from the control. So the control would be that G F P just being produced in the cell. And then you say, well, if we have the G F P expression being controlled by the CIS elements, do we see the fluorescence being higher or lower than the control? And that will give us an idea of what the cis regulatory elements, what their effect is on gene expression. So again, which of the following techniques used to study the effects of these elements on post transcription and gene regulation choice. C A reporter gene assay. See you in the next video.

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Key Concepts

Cis-Elements in mRNA

Reporter Gene Assays Using GFP

Posttranscriptional Gene Regulation Mechanisms