RNA

Part:BBa_K115002:Experience

Designed by: Bastiaan van den Berg   Group: iGEM08_TUDelft   (2008-07-15)
Revision as of 18:23, 28 October 2010 by Jim6904 (Talk | contribs) (→‎Applications of BBa_K115002)

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Applications of BBa_K115002

The parts were tested in luciferase constructs, by measuring luciferase activity after growing at certain temperatures. The fold increase of luciferase activity between cultures of cells grown at 20ºC and 37ºC is displayed in figure 1.

In this figure, 12 (the leftmost) is construct K115012, the reference strain, which has no thermosensitive RNA, but just B0032 as ribosome binding site. This is to compare for normal temperature induced behaviour in cells. The other constructs from 29-36 are K115029 to K115036, please look at the experience tab of these constructs to see which thermometer RNA they contain.

Figure 1. Fold increase of luminescence per ug of total protein of each construct in respect to luminescence measured at 20ºC. Numbers in the legend represent the last two numbers of the construct name, e.g. 12 = BBa_K115012. Four samples were measured in duplo for every data point. Error bars represent two times SEM.


For more documentation, visit our wiki [http://2008.igem.org/Team:TUDelft (link)].


2010IGEM-TEAM NCTU_formosa's application

We using this parts as a regulator in our project's low-temperature release system.

We constructed series circuits' test as following article.


First, we made A+B circuit in the PSB3K3 plasmid. in this way we can use it to test by Using Flow Cytometry to obtain the data of A+B (for40°C, 37°C, 30°C, and 25°C)

TPcon1.jpg

and the following figures is our testing results. To test the efficiency of the temperature induced RBS and the interaction between tetR gene and ptet promoter, we replaced the crystal protein gene with GFP (green fluorescence protein) gene. In this scenario, GFP will simulate the crystal protein’s production, as it is easily detected by flow cytometry.

TECON F4.jpg

To conclude, we have verified that our strategy works. It demonstrates the regulation of GFP in a temperature dependent fashion, as it is evident the mean GFP values are significantly lower at T>37° C.

We expect our circuit design to allow steady bacteria growth and inhibit crystal protein at T>37° C, and high protein production and low bacteria growth rate at T<37° C

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