Part:BBa_K1140006:Experience

Applications of BBa_K1140006

The team observed that this part works correctly in E. coli K12.

Figure 1. Temperature dependence of mCherry translation by u6 RNA thermometer in E. coli. Tet repressor is NOT present in this test. Two control cultures without mCherry sequence are included for growth and color comparison. a) Control 30ºC b)Control 37ºC c)30ºC d)37ºC.

Figure 2. Relative fluorescence under 25, 30, 37 and 42 celcius grades in E. coli. M1 is a group of cultures used by UANL_Mty-Mexico team. Tet repressor is NOT present in this test.

Surprisingly, we obtained different behaviors in clones transformed with the same DNA (figure 3). All measurements were performed at least in triplicate, the aritmethic mean is shown. Figure 2 shows the behavior of our best clone, dubbed M1. M12 clone, showing a weird behavior, is to be sequenced to verify if this outcome is due to mutation or intrinsic cellular noise.

Figure 3. Behavior of different clones transformed with this construction (M1, 2, 11 and 12). Relative fluorescence under 25, 30, 37 and 42 celcius grades in E. coli.

Mathematically, we found that a simple gaussian function fits our data well, and it provides us a way to quantify the strength (amplitude), optimal value (horizontal shift), and definition or clearness (width) of our RNAT activity (figure 4). We believe positive slope is due to RNAT melting, while negative slope is due to increase in the overall protein degradation rate due to higher temperatures. This function also allows for comparisons between different RNAT, as well as being potentially predictive for non verified temperatures.

Figure 4. Gaussian Function fitting of the experimental data shown in figure 3.

User Reviews

UNIQ2cd4dce1750d0e97-partinfo-00000000-QINU UNIQ2cd4dce1750d0e97-partinfo-00000001-QINU