Part:BBa_K676013
Mu/Spy Ligation
A device to see the difference in the expression of GFP between the presence and absence of a gyrase binding site on a plasmid molecule.
Usage and Biology
The Mu phage GBS which was ligated to the Spy promoter device (Part:BBa_K239009) and it displayed a higher expression of GFP in vivo compared to the old Spy promoter device without the Mu phage GBS. The higher expression level may possibly be due to the fact that negative supercoiling in the DNA promote easier transition from the close promoter complex to open promoter complex for higher rate of transcription. The higher level of expression therefore indicates that the introduction of such GBS into plasmids carrying DNA vaccine or therapeutic genes will be able to trigger a better immune response and hence induce a fully developed protection against the target infection. Although this is only a small experiment to confirm the higher expression, further studies are required to verify it.
The higher level of expression can be used to make a more sensitive Spy promoter device (shearometer) in order to detect lower shear stress levels in the cells during fermentation and downstream processing.
As with the Mu phage GBS (Part:BBa_K676001) ligated to the Spy promoter device, we also observed a slightly higher supercoiling efficiency (about 10%) and better uniformity (smaller range of supercoiled topoisomers) in the mini-prep plasmid sample when compared to the control plasmid (Spy promoter device with no GBS).
Sequence and Features
- 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 191
- 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI.rc site found at 1203
| n/a | Mu/Spy Ligation |