Difference between revisions of "Part:BBa K1017603"

Revision as of 17:17, 22 September 2013

37℃ RBS + LuxR

This unit includes 37℃ RBS and LuxR. 37℃ RBS, a RBS which on/off is controlled by temperature. LuxR, a protein which can activate lux promoter after binding with AHL. This unit act as a biotheometer, producing lux protein when the environment is higher than 37℃ and stop expressing when lower than 37℃.

In our project, we use this part as a biotheometer to sense the environment. When the temperature is higher than 37℃, the downstream gene LuxR is expressed. Therefore , P_lux is activated and the downstream gene sRNA is produced, causing the stop of target region translate. When lower than 37℃, P_lux is no longer activated , which means sRNA production shuts down. Thus, the target region can be expressed normally.

Mechanism

When the temperature is higher than 37℃, 37℃ RBS functions normally and the downstream gene can be expressed. In contrast with high temperature,37℃ RBS forms a hairpin structure that block the ribosome binding when lower 37℃. Thus, the downstream gene, mGFP, is not able to translate.

Fig 1. When the temperature increases, the secondary structure near the SD site would change its conformation and allow the ribosome to bind to initiate translation

Data

From chart 1, we can clearly found that the time to reach same O.D. value(0.8) various as the temperature changes. The expression of mGFP under 37°C and 42°C reached a higher OD value in less time than the expression below 37°C. This demonstrates the fact that 37°C RBS functions more efficiently above or at 37°C. On the other hand, the fact that 37°C RBS leads to moderate expression even under 37°C degree might be a problem, since then we cannot make sure that only single gene is expressed under 37°C.

Chart 1. 37°C RBS efficiency under different temperatures.

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
COMPATIBLE WITH RFC[12]
21
COMPATIBLE WITH RFC[21]
23
COMPATIBLE WITH RFC[23]
25
COMPATIBLE WITH RFC[25]
1000
COMPATIBLE WITH RFC[1000]

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 In our project, we use this part as a biotheometer to sense the environment. When the temperature is higher than 37℃, the downstream gene LuxR is expressed. Therefore , P<sub>lux</sub> is activated and the downstream gene sRNA is produced, causing the stop of target region translate. When lower than 37℃, P<sub>lux</sub> is no longer activated , which means sRNA production shuts down. Thus, the target region can be expressed normally.
+== Mechanism ==
 <br>When the temperature is higher than 37℃, 37℃ RBS functions normally and the downstream gene can be expressed. In contrast with high temperature,37℃ RBS forms a hairpin structure that block the ribosome binding when lower 37℃. Thus, the downstream gene, mGFP, is not able to translate.[[File:NCTU RNAthermometer.jpg|center|Fig 1. When the temperature increases, the secondary structure near the SD site would change its conformation and allow the ribosome to bind to initiate translation]]
-<br>From chart 1., we can clearly found that the time to reach same O.D. value(0.8) various as the temperature changes. The expression of mGFP under 37°C and 42°C reached a higher OD value in less time than the expression below 37°C. This demonstrates the fact that 37°C RBS functions more efficiently above or at 37°C. On the other hand, the fact that 37°C RBS leads to moderate expression even under 37°C degree might be a problem, since then we cannot make sure that only single gene is expressed under 37°C.
+== Data ==
+<br>From chart 1, we can clearly found that the time to reach same O.D. value(0.8) various as the temperature changes. The expression of mGFP under 37°C and 42°C reached a higher OD value in less time than the expression below 37°C. This demonstrates the fact that 37°C RBS functions more efficiently above or at 37°C. On the other hand, the fact that 37°C RBS leads to moderate expression even under 37°C degree might be a problem, since then we cannot make sure that only single gene is expressed under 37°C.
 [[File:NCTU 37RBS result.png|center|600px|Chart 1. 37°C RBS efficiency under different temperatures.]]