Difference between revisions of "Part:BBa J102005"

Latest revision as of 02:26, 25 August 2010

Biological Capacitor

This is a biological capacitor, where the output of the device is proportional to the temporal derivative of the concentration of the input. If one considers current to be PoPS, and voltage to be the concentration of the input molecule (where ground is 0M), this is analogous to an electrical capacitor. In this case, the input is anhydrotetracycline, and the output is GFP (with an LVA tag). A modular version of this device, where the user can define his or her own input and output, is also available (BBa_J102003).

The concentration of AHL, a small molecule which activates luxR, is used as a chemical dial to control the transcription rate from J102001. For the capacitor to display the desired behaviour, the promoters must be of approximately equal strength. The more AHL in the medium, the more transcription occurs from J102001, the reverse lux promoter. In order to tune the capacitor, grow bacteria containing the plasmid in media with different concentrations of AHL, and test for the desired behaviour.

Please refer to the experience section for extensive modelling data.

How it Works:

-TetR binds to the tet promoter, inhibiting transcription of downstream genes.

-ATc inhibits this repression by binding to TetR.

-LuxR, a transcriptional activator, up-regulates transcription from the lux promoter in the presence of the signalling molecule AHL. Thus, AHL is used as a chemical dial to control device behaviour.

-Modelling data showed that the Lux promoter must be stronger than the Tet promoter for the device to function properly. We use AHL to tune the relative strength of the promoters.

-The transcript produced by the Lux promoter is complementary to the GFP mRNA. The mRNA and the reverse transcript anneal, prohibiting translation of the GFP, but allowing further translation of the LuxR mRNA. This is the reason for basal levels of GFP expression given any constant level of ATc.

-When the ATc concentration is increased or decreased, GFP and LuxR production quickly follow. However, there is a time lag before the production of the reverse transcript reaches equilibrium, allowing more GFP to be translated. This time lag is the cause of the capacitive behaviour observed.

Please contact Michael Pettigrew with any questions regarding this part.

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
INCOMPATIBLE WITH RFC[12]
Illegal NheI site found at 7
Illegal NheI site found at 30
21
COMPATIBLE WITH RFC[21]
23
COMPATIBLE WITH RFC[23]
25
COMPATIBLE WITH RFC[25]
1000
INCOMPATIBLE WITH RFC[1000]
Illegal BsaI.rc site found at 1615

@@ Line 4: / Line 4: @@
 [[image:Capacitorequation.JPG]]
-This is a biological capacitor, where the output of the device is proportional to the temporal derivative of the concentration of the input. If one considers current to be PoPS, and voltage to be the concentration of the input molecule (where ground is 0M), this is analogous to an electrical capacitor. In this case, the input is anhydrotetracycline, and the output is GFP (with an LVA tag). A modular version of this device, where the user can define his or her own input and output, is also available (BBa_J102003).
+This is a biological capacitor, where the output of the device is proportional to the temporal derivative of the concentration of the input. If one considers current to be PoPS, and voltage to be the concentration of the input molecule (where ground is 0M), this is analogous to an electrical capacitor. In this case, the input is anhydrotetracycline, and the output is GFP (with an LVA tag). A [https://parts.igem.org/Part:BBa_J102003 modular version] of this device, where the user can define his or her own input and output, is also available (BBa_J102003).
 The concentration of AHL, a small molecule which activates luxR, is used as a chemical dial to control the transcription rate from J102001. For the capacitor to display the desired behaviour, the promoters must be of approximately equal strength. The more AHL in the medium, the more transcription occurs from J102001, the reverse lux promoter. In order to tune the capacitor, grow bacteria containing the plasmid in media with different concentrations of AHL, and test for the desired behaviour.
+'''Please refer to the experience section for extensive modelling data.'''
 [[image:Capacitorcircuit.JPG]]
@@ Line 14: / Line 16: @@
 -TetR binds to the tet promoter, inhibiting transcription of downstream genes.
--ATc inhibits this repression.
+-ATc inhibits this repression by binding to TetR.
 -LuxR, a transcriptional activator, up-regulates transcription from the lux promoter in the presence of the signalling molecule AHL. Thus, AHL is used as a chemical dial to control device behaviour.
--The transcript produced by the lux promoter is complementary to the GFP mRNA. The mRNA and the reverse transcript anneal, prohibiting translation of the GFP, but allowing further translation of the LuxR mRNA.
+-Modelling data showed that the Lux promoter must be stronger than the Tet promoter for the device to function properly. We use AHL to tune the relative strength of the promoters.
+-The transcript produced by the Lux promoter is complementary to the GFP mRNA. The mRNA and the reverse transcript anneal, prohibiting translation of the GFP, but allowing further translation of the LuxR mRNA. This is the reason for basal levels of GFP expression given any constant level of ATc.
 -When the ATc concentration is increased or decreased, GFP and LuxR production quickly follow. However, there is a time lag before the production of the reverse transcript reaches equilibrium, allowing more GFP to be translated. This time lag is the cause of the capacitive behaviour observed.
+'''Please contact [[User:pettigr7|Michael Pettigrew]] with any questions regarding this part.'''