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Team:Washington/Future
From 2009.igem.org
(Difference between revisions)
| Line 5: | Line 5: | ||
# Attempt to add additional proteins into the vector and test for functionality | # Attempt to add additional proteins into the vector and test for functionality | ||
# Vary linker lengths | # Vary linker lengths | ||
| - | # Make a simpler version for trouble shooting the secretion | + | # Make a simpler version for trouble shooting the secretion system: [6x-His]-[NheI]-[prtB] |
# Transfer to a vector with the same origin and resistance as described in the original secretion system (pBR322+Carb). | # Transfer to a vector with the same origin and resistance as described in the original secretion system (pBR322+Carb). | ||
# Add a lacI into the expression and target vectors so repression is hard-coded into the vector and expression can be induced regardless of the cell line. | # Add a lacI into the expression and target vectors so repression is hard-coded into the vector and expression can be induced regardless of the cell line. | ||
| Line 14: | Line 14: | ||
# Add original upstream DNA (50bp) before the native RBS to ensure proper function | # Add original upstream DNA (50bp) before the native RBS to ensure proper function | ||
# Add an arabanose inducible promoter for better control over secretion system activation | # Add an arabanose inducible promoter for better control over secretion system activation | ||
| - | # Combine with target vector so entire secretion system | + | # Combine with target vector so entire secretion system is contained in one plasmid. |
Revision as of 05:15, 15 October 2009
- Target Construct
- Attempt to add additional proteins into the vector and test for functionality
- Vary linker lengths
- Make a simpler version for trouble shooting the secretion system: [6x-His]-[NheI]-[prtB]
- Transfer to a vector with the same origin and resistance as described in the original secretion system (pBR322+Carb).
- Add a lacI into the expression and target vectors so repression is hard-coded into the vector and expression can be induced regardless of the cell line.
- Secretion System
- Transfer to a Chlor resistance, p15A origin vector as used in the original papers
- Add original upstream DNA (50bp) before the native RBS to ensure proper function
- Add an arabanose inducible promoter for better control over secretion system activation
- Combine with target vector so entire secretion system is contained in one plasmid.
- Display System
- Test additional proteins in the new custom display construct (GFP, OpdA, etc)
- Test designed proteins from FoldIt
