From 2007.igem.org

Edinburgh > Yoghurt

Introduction | Applications | Design | Status | References

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Contents 1 Status 1.1 Vanillin Genes 1.1.1 Sam5 & Sam8 1.1.2 COMT 1.1.3 fcs & ech 1.1.4 fcs 1.1.5 ech 1.2 vector 1.3 Enzyme Accession Numbers

Status

Vanillin Genes

Sam5 & Sam8

• Saccharothrix espeanensis has arrived from DSMZ culture collection

requires reviving and fcs and ech genes being PCR'd out

• Sam8 and Sam5 expression vector kindly donated by Dr Anton Linnenbrink from the Institute for Pharmazeutische Wissenschaften in Germany

vector has been transformed into E. coli

need to PCR out genes when primers arrive

COMT

• have grown up alfalfa seeds and genomic plant DNA has been extracted

attempted to amplify up COMT via PCR, but have had no promising results so far

• RNA has been isolated from alfalfa

have attempted to amplify up cDNA of COMT gene and are waiting on results

fcs & ech

• Pseudomonas fluorescens has arrived from NCIMB culture collection and has been revived

fcs

• have carried out several PCRs to amplify out the fcs gene from Pseudomonas fluorescenes

PCRs did not result in one PCR product being produced, problem is most probably due to the P. fluorescens genome being of high GC content

are attempting to purify out fcs gene from agrose gel of PCR products

ech

• the first set of primers ordered for ech did not work, as no PCR product was present after several PCR reactions

• have sent away for a second set of primers and are awaiting their arrival

vector

• we are going to modify a vector kindly donated by Norwich University, which can act as a shuttle vector between E. coli and a broad range of gram positive hosts, including Bacillis subtillis, Lactobacillus and Lactococcus

• firstly we will ligate MITs wintergreen scent generating device into the vector to characterise how the vector works in our chosen gram positive organisms
• secondly, when we have a working vanillin synthesis pathway, the vanillin biosynthesis pathway will be inserted into the vector and then transformed into Lactobacillus under the control of the arabinose promoter

Lactobacillus may then be used to make yoghurt giving us self flavouring yoghurt, when the arabinose inducer is added

Enzyme Accession Numbers

Sam5 and Sam8 Accession: DQ357071

Saccharothrix espanaensis culture collection: DSM 44229

COMT Accession: M63853

fcs and ech Accession: DQ119298

Pseudomonas fluorescens culture collection: NCIMB 9046

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Introduction | Applications | Design | Status | References