Difference between revisions of "Part:BBa K817002"
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It's a fatty acyl-CoA responsive promoter which will express the downstream gene in the presence of fatty acid. | It's a fatty acyl-CoA responsive promoter which will express the downstream gene in the presence of fatty acid. | ||
| − | < | + | ===Method=== |
| − | === | + | To evaluate the P<sub>fadBA</sub> promoter function, we design a P<sub>fadBA</sub>-mRFP reporter construct. In experimental group we add oleic acid in media and compared to control group, which is the same colony and identical amount of bacteria but without addition of oleic acid. After 5 hr induction we measure the man fluorescence intensity to get the expression level of P<sub>fadBA</sub> promoter in either group. |
| + | |||
| + | ===Protocol=== | ||
| + | #10 μL bacterial culture was cultured in 5 mL LB at 37。C, with suitable concentration of antibiotics shaking for 18 hr. | ||
| + | #For experimental group, 480 μL of bacteria culture was transferred to 1.5 mL eppendorf tube and added with 20 μL of oleic acid and IGEPAL(detergent) in. For control group, 500 μL bacterial culture was added in 1.5 mL eppendorf tube. | ||
| + | #The tubes were incubated at 37℃ 5 hr for induction. | ||
| + | #100 μL of bacterial culture was transferred tino 96-well plate. The mRFP fluorescence intensity was measured (Excitaion: 580 nm, Emmision: 610 nm). | ||
| + | |||
| + | ===Data=== | ||
| + | [[FIle:NTU-Taida-Result-PfadBA-fig1.png|450px|thumb|center|TEST]] | ||
| + | '''Oleic acid induction group shows higher expression of mRFP.''' | ||
| + | |||
| + | ===Conclusion=== | ||
| + | Our P<sub>fadBA</sub> promoter can work in response to environmental fatty acid change, which acts as an important sensor for our bacterial device – secreting GLP-1 when host is fed with fatty diet. | ||
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<span class='h3bb'>Sequence and Features</span> | <span class='h3bb'>Sequence and Features</span> | ||
<partinfo>BBa_K817002 SequenceAndFeatures</partinfo> | <partinfo>BBa_K817002 SequenceAndFeatures</partinfo> | ||
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<!-- Uncomment this to enable Functional Parameter display | <!-- Uncomment this to enable Functional Parameter display | ||
Revision as of 11:45, 5 October 2012
PfadBA
It's a fatty acyl-CoA responsive promoter which will express the downstream gene in the presence of fatty acid.
Method
To evaluate the PfadBA promoter function, we design a PfadBA-mRFP reporter construct. In experimental group we add oleic acid in media and compared to control group, which is the same colony and identical amount of bacteria but without addition of oleic acid. After 5 hr induction we measure the man fluorescence intensity to get the expression level of PfadBA promoter in either group.
Protocol
- 10 μL bacterial culture was cultured in 5 mL LB at 37。C, with suitable concentration of antibiotics shaking for 18 hr.
- For experimental group, 480 μL of bacteria culture was transferred to 1.5 mL eppendorf tube and added with 20 μL of oleic acid and IGEPAL(detergent) in. For control group, 500 μL bacterial culture was added in 1.5 mL eppendorf tube.
- The tubes were incubated at 37℃ 5 hr for induction.
- 100 μL of bacterial culture was transferred tino 96-well plate. The mRFP fluorescence intensity was measured (Excitaion: 580 nm, Emmision: 610 nm).
Data
Oleic acid induction group shows higher expression of mRFP.
Conclusion
Our PfadBA promoter can work in response to environmental fatty acid change, which acts as an important sensor for our bacterial device – secreting GLP-1 when host is fed with fatty diet.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]