Difference between revisions of "Part:BBa K1017602"

Revision as of 12:05, 22 September 2013

37℃ RBS + mGFP

This biobrick is comprised of 37 degrees Celsius RBS and mGFP. 37℃ RBS, a RBS which on/off is controlled by temperature. mRFP is a red fluorescent protein. In our project, we use this part to conduct the test of temperature regulated-system. By adding a P_cons promoter at the upstream gene , mRFP is constantly expressing and produce red fluorescent protein. Then measure the O.D. value to determine the level of expression at different temperature.

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

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
INCOMPATIBLE WITH RFC[1000]
Illegal BsaI.rc site found at 702

@@ Line 2: / Line 2: @@
 <partinfo>BBa_K1017602 short</partinfo>
-This biobrick is comprised of  37 degrees Celsius RBS and mGFP. 37℃ RBS, a RBS which on/off is controlled by temperature. mRFP is a red fluorescent protein. <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]]
+This biobrick is comprised of  37 degrees Celsius RBS and mGFP. 37℃ RBS, a RBS which on/off is controlled by temperature. mRFP is a red fluorescent protein.
+In our project, we use this part to conduct the test of temperature regulated-system. By adding a P<sub>cons</sub> promoter at the upstream gene , mRFP is constantly expressing and produce red fluorescent protein. Then measure the O.D. value to determine the level of expression at different temperature.
+<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.
+<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.]]