IGEM:MIT/2006/Notebook/2006-6-7: Difference between revisions
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*Restriction sites | *Restriction sites | ||
**JAMT- None. | **JAMT- None. | ||
**SAMT (A. Majus and C. Breweri) - None | |||
*Codon optimization | *Codon optimization | ||
**JAMT- Already transferred to ''E. coli'' and ''S. cerevisiae'' | **JAMT- Already transferred to ''E. coli'' and ''S. cerevisiae'' | ||
**SAMT - (A. Majus and C. Breweri) - Already ported to e. coli | |||
*Registry accounts | *Registry accounts | ||
**Stephen- Yes | **Stephen- Yes | ||
**Kate- Yes | |||
*Install DNA 2.0 software? | *Install DNA 2.0 software? | ||
**Stephen- Yes | **Stephen- Yes | ||
**Kate - Yes (although have not used) | |||
==Systems & Applications== | ==Systems & Applications== |
Revision as of 07:40, 8 June 2006
Compound Database
Transport
- SA can be transported into/out of tobacco cells (http://ejournal.sinica.edu.tw/bbas/content/1999/4/bot404-03.html)
- BA was added exogenously, and methyl benzoate (MB) was produced, so BA can enter E. coli. MB concentrations were measured without needing to lyse the cells, so MB can exit the cell. (Reference; it is the first paper under this section)
- In an experiment done by Horton et al., isoamyl alcohol was added exogenously and was taken up by the E. coli. Concentration of the product, isoamyl acetate, was then measured without lysing the cells, so we know that the product will be excreted. (see Horton et al. paper in this section.)
Biosynthesis
salicylic acid/benzoic acid
http://www.hort.purdue.edu/rhodcv/hort640c/secprod/sec7.gif
jasmonic acid
- Many pathways described at http://www.pubmedcentral.nih.gov/picrender.fcgi?artid=1066929&blobtype=pdf
Toxicity
- Immediately following exposure of E. coli to benzoate (20 mM = 2.42μg/mL), the cell doubling rate reduced 10-fold (this is at pH 6.5; at pH 8.0, there is no significant change in doubling rate). But we have to keep in mind that benzoate was not being metabolized, so benzoate was simply accumulating in the cell, affecting the pH (this is what was being studied, actually). Methyl benzoate, since it is uncharged, may not have this same effect.
- Lambert LA, Abshire K, Blankenhorn D, and Slonczewski JL. Proteins induced in Escherichia coli by benzoic acid. J Bacteriol. 1997 Dec;179(23):7595-9. DOI:10.1128/jb.179.23.7595-7599.1997 |
Concentration
What ranges of concentrations should we be adding?
banana
- 10 mM isoamyl alcohol
jasmine
- 1 mM JA and 1 mM S-adenosyl methionine (SAM)
salicylic acid or benzoic acid
- SAMT (A. Majus and C. breweri) 5 μg/ml of SA (or BA)
Sequence analysis
- Restriction sites
- JAMT- None.
- SAMT (A. Majus and C. Breweri) - None
- Codon optimization
- JAMT- Already transferred to E. coli and S. cerevisiae
- SAMT - (A. Majus and C. Breweri) - Already ported to e. coli
- Registry accounts
- Stephen- Yes
- Kate- Yes
- Install DNA 2.0 software?
- Stephen- Yes
- Kate - Yes (although have not used)
Systems & Applications
Things to get
- Precursor molecules etc.
- BL21(DE3)+pLysS competent cells (the Endy Lab has these)
- Arab. JMT
Parts in the registry
<bbpart>BBa_J45000</bbpart> <bbpart>BBa_J45001</bbpart> <bbpart>BBa_J45002</bbpart> <bbpart>BBa_J45003</bbpart> <bbpart>BBa_J45004</bbpart> <bbpart>BBa_J45005</bbpart> <bbpart>BBa_J45006</bbpart> <bbpart>BBa_J45007</bbpart> <bbpart>BBa_J45011</bbpart>