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Sometimes in synthetic biology, we need to know rates of transcription of one promoter in relation to others (particularly inducible vs constitutive) in order to perform tasks like balancing transcription products and ensuring an excess of one product over another. As far as I know, there is no standard method of characterizing promoter strengths in relation to one another. However, a logical unit for transcription rate made (by organism or in vitro) could look something like:

Promoter Strength = number of transcript made / (concentration of RNA polymerase) (length of transcript) (unit of time).

Why hasn't this been done yet? What other factors could contribute to rate of expression in a particular organism? Is there another system for comparing promoter strengths?

Note: a similar system could be used for strength of repression.

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What if there are multiple regulatory elements acting on a given locus? –  Amory Jul 30 '13 at 15:57
    
I think the quantity you suggest is suitable only if the relations between the different factors is linear. Is there any proof of that? –  Bitwise Jul 30 '13 at 20:19
    
@Amory, this is exactly why a characterized number would be useful to define promoter strength. You would be able to quantify a particular promoter's strength with or without activators/repressors, as well as in relation to other promoters with their respective regulatory elements. –  LanceLafontaine Jul 30 '13 at 23:29
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The destruction ratio of the mRNA also varies, for example. Also, different mRNAs may have different affinity to the ribosomes (for instance, differences in Shine-Dalgarno sequence in prokaryots). Finally, unless you're using retrotranscriptase RT-qPCR, you shoudn't have a fiable method to quantify gene expression. –  Miguel Ángel Naranjo Ortiz Aug 29 '13 at 22:44
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3 Answers 3

Transcription rates are very context dependent. This shouldn't surprise you, since the same genes aren't expressed in your lungs and your eyes. They're also very environment dependent, this shouldn't surprise you either, since it's pretty obvious every organism is going to respond to temperature, their current nutritional state and so on.

So, in order to define promoter strength, you'd first need to define the exact conditions under which you're measuring it, including the exact strain you're measuring it in. Because if you get any of those even slightly different you're going to get a different answer. Doing this might get you a consistent number but you're abstracting away the most important parts of differences in transcription anyway: how it varies between cells and contexts.

Moreover, even if got past these hurdles you'd not have a particularly useful answer anyway since the rate of transcription is only a small part of the whole process. Why not consider mRNA durability, or rates of translation, or protein durability? What matters very much depends on what you're considering.

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I do realize that transcription is very context-dependent, but disagree about its importance. Even if strictly used in characterized in vitro systems, a number associated with a promoter is very useful, and can be extrapolated to its general abilities for transcription in vivo. –  LanceLafontaine Jul 30 '13 at 23:26
    
No, it can't. Consider DUO1 in Arabidopsis: it's a gene that expresses strongly in the germline but is entirely unexpressed in somatic tissue (and, in fact, is detrimental if it is). How will expression in vitro be extrapolated then? If your in vitro model is similar to the germline you'll get results irrelevant to any other cell; if your results are relevant to the somatic tissue you'll fail to understand key processes in the germline. The major issue in gene expression is its variation. –  Jack Aidley Jul 30 '13 at 23:37
    
Not all genes need to be expressed in their "proper" context. Think of the Tet and lambda pR promoters being expressed in E coli (or really synthetic biology as a whole). If the purpose is exploiting or engineering and not discovering novel protein functions at a particular developmental stage, these promoters strengths are extremely relevent. –  LanceLafontaine Jul 30 '13 at 23:46
    
the measurement - counting # of mRNA from a specific cell state over a period of time would be difficult to do. a single cell measurement where you are simultaneously monitoring the promotion state of the gene. Sounds like quite a bit of work! –  shigeta Jul 31 '13 at 5:58
    
@LanceLafontaine: Extremely relevant for a particular purpose carried out in a particular way. That doesn't call for a standard measure; instead if you want to compare the effectiveness of two promoters you can simply compare them by Western Blotting or qPCR and see what you find. If you are desperate for a number, this will quantify the difference. –  Jack Aidley Jul 31 '13 at 10:39
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It's true that the expression of a promoter is context-dependent, i.e. depends on the overall status of the cell, but some work has been made to boil down such context-dependency into some measurable parameter. For example, for constitutive promoters such a parameter seems to be the growth rate. You could characterize promoters by that:

Gerosa, L. et al. Dissecting specific and global transcriptional regulation of bacterial gene expression. Mol. Syst. Biol. 9, 658 (2013)

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Years of discussion about this:

"Polymerases Per Second.

A standard unit to measure the inputs and outputs of a BioBrick Device.

PoPS have yet to be directly measured in vivo."

http://openwetware.org/wiki/PoPS

*understanding this would be a great feat and it would get us one step closer to measuring complex systems.

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