Difference between revisions of "Part:BBa K2379007"

Latest revision as of 03:26, 2 November 2017

pH sensitive system with RNA pHmeter

The RNA pHmeter BBa_K2379012(BBa_K2379012) is expressed under the regulation of the pLAC promoter(BBa_R0011). Following the RNA pHmeter, the construct contains GFP (BBa_E0040) as the reporter which is followed by the double terminator (BBa_B0014). Expression of GFP is regulated by the RNA pHmeter and hence will be produced only at pH 8.5

Usage and Biology

ABOUT THIS PART :

We used this part to regulate gene expression at high pH ,i.e, at pH 8.5. This part functions as a pH-sensitive RNA element which contains the SraF gene. This variant of the part is one with the bio-brick scar site. This part works as expected and when GFP is placed downstream of this gene, maximum fluorescence was observed at pH 8.5.

CHARACTERISATION STUDIES FOR THIS PART:

1. The above graph indicates the fluorescence expression at three different time periods (each recorded at an interval of 60 minutes)

2. The cell cultures were grown at three different pH : 7.0 ,8.0 and 8.5. Fluorescence values were recorded at excitation and emission peaks of 460nm and 515nm respectively.

3. Normalised fluorescence was calculated by dividing observed fluorescence by cell density at OD600.

4. Duplets were used for characterisation and the error bars indicate the Standard Deviation of the mean values.

CONCLUSION:

Expected results were obtained for the RNA pHmeter containing biobrick scar site. There was only an average 22.06% increase in the expression of GFP from pH 7 to pH 8.5 at all the three time points which is comparable to that obtained for RNA pHmeter with wild scar. This proves that the changes made in the gene sequence hasn’t affected the way in which the part functions. Though the expression here is controlled by both the promoter and the SraF gene, the latter has more ascendancy which is evident from the maximum fluorescence observed at pH 8.5 irrespective of the time points at which it is recorded. Thus the RNA pHmeter with biobrick scar works as expected.

Sequence and Features