Part:BBa_K3151029
yhjH + Riboswitch
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 1362
- 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI.rc site found at 1344
Overview
The activity of the phosphodiesterase determines the concentration of the cyclic-di-GMP that is bound to the terminator stem of the riboswitch. This altogether then determines the production of the green fluorescent protein. The chief hypothesis formed was that the intracellular concentration of cyclic-di-GMP does not affect the activity of the riboswitch with no terminator stem loop (RON) since there is no component to stop the transcription of the fluorescent protein. However, it does affect the Riboswitch with the terminator stem loop (ROFF). As seen in (Fig. 1) when YHJH (Phosphodiesterase) was induced and was activated, its activity had a decreasing effect on cyclic-di-GMP, hence, increasing the production of eGFP.
Assembly and Design
The parts were digested using enzymes EcoRI and SpeI and then ligated using standard assembly,
Characterization
Figure 1. Agarose gel of sequence confirmed parts, showing the size (bp) of R OFF with YHJH.
Figure 2.The constitutive riboswitch was expressed in DH5ɑ cells. The black lines represent RON samples, the blue lines represent ROFF samples. All RON samples were observed to express more Green Fluorescent Protein (GFP) than corresponding ROFF samples. The effect of sucrose is shown as inhibiting the expression of GFP, shown in the decrease of lower.
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