Team:KULeuven/Lab/Vanillin Production
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(New page: =VANILLIN SYNTHESIS= ==Goal== Making vanillin from tyrosine in a five-step pathway. Making vanillin from tyrosine ==requ...) |
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=VANILLIN SYNTHESIS= | =VANILLIN SYNTHESIS= | ||
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[[Image:Vanillin_Biosynthesis_Pathway.jpg|center|thumb|365px|Making vanillin from tyrosine]] | [[Image:Vanillin_Biosynthesis_Pathway.jpg|center|thumb|365px|Making vanillin from tyrosine]] | ||
- | == | + | ==Required== |
- | + | Biobricks: | |
- | + | *SAM8: {{kulpart|BBa_I742142}}: coding sequence without RBS | |
- | + | *SAM5: {{kulpart|BBa_K238007}}: RBS site + PstI restriction site removed | |
- | + | *COMT: {{kulpart|BBa_I742107}}: coding sequence without RBS | |
- | + | *FCS: {{kulpart|BBa_I742115}}: RBS + fcs in pSB1A2 | |
- | + | *ECH: {{kulpart|BBa_I742113}}: RBS + ech in pSB1A2 | |
==Where from== | ==Where from== | ||
+ | * SAM8, SAM5, COMT, FCS, ECH will be sent to us from the French Lab from the university Edinburgh | ||
+ | |||
+ | ==Steps== | ||
+ | *We will be working in three different stages | ||
+ | #Testing the transformation of tyrosin to ferrulic acid. The enzymes are put under a constitutive promoter and transcribed. Ferrulic acid concentrations are then measured with GC | ||
+ | #Testing the transformation of ferrulic acid to vanillin. The enzymes are put under a constitutive promoter and transcribed. Ferrulic acid concentrations are then measured with GC | ||
+ | #If both are tested and ok, they can be combined into one plasmid and again tested. | ||
+ | #Then, the locks need to be added. | ||
- | * | + | *Testing the enzymes by detecting the endproducts through gaschromatography. |
- | + | *One problem might be that the tyrosine of E. coli is depleted too fast | |
- | * | + |
Latest revision as of 06:25, 18 August 2009
Contents |
VANILLIN SYNTHESIS
Goal
Making vanillin from tyrosine in a five-step pathway.
Required
Biobricks:
- SAM8: : coding sequence without RBS
- SAM5: : RBS site + PstI restriction site removed
- COMT: : coding sequence without RBS
- FCS: : RBS + fcs in pSB1A2
- ECH: : RBS + ech in pSB1A2
Where from
- SAM8, SAM5, COMT, FCS, ECH will be sent to us from the French Lab from the university Edinburgh
Steps
- We will be working in three different stages
- Testing the transformation of tyrosin to ferrulic acid. The enzymes are put under a constitutive promoter and transcribed. Ferrulic acid concentrations are then measured with GC
- Testing the transformation of ferrulic acid to vanillin. The enzymes are put under a constitutive promoter and transcribed. Ferrulic acid concentrations are then measured with GC
- If both are tested and ok, they can be combined into one plasmid and again tested.
- Then, the locks need to be added.
- Testing the enzymes by detecting the endproducts through gaschromatography.
- One problem might be that the tyrosine of E. coli is depleted too fast