Team:Washington/Accomplishments

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Accomplishments & Submitted BioBricks

Accomplished

Create Protein Expression Vector for IPP system
- [http://partsregistry.org/wiki/index.php/Part:BBa_K215000 BBa_K215000], Strong laq promoter with strong RBS
Demonstrate Stable, Functional, His-Tagged Fusion protein from expression vector
- [1]See Project Page: Target
Synthesize and BioBrick type I secretion system from Erwinia Chrysanthemi
- [http://partsregistry.org/wiki/index.php/Part:BBa_K215107 BBa_K215107], prtDEF from Erwinia Chrysanthemi under constitutive expression in low copy plasmid
Develop assay for looking for secreted proteins
- [2]See Project Page: Secretion
Extensively characterize Legacy Surface Display BioBricks in registry[3]
- [http://partsregistry.org/wiki/index.php/Part:BBa_J36848 BBa_J36848]
- [http://partsregistry.org/wiki/index.php/Part:BBa_J36849 BBa_J36849]
- [http://partsregistry.org/wiki/index.php/Part:BBa_J36850 BBa_J36850]
- [http://partsregistry.org/wiki/index.php/Part:BBa_J36851 BBa_J36851]
Develop new modular surface display system
Computationally design novel Biotin-binding protein using Rosetta
Submit at least one new Biobrick functioning as designed
- Target Expression Vector
- OPDA [4]
Gain real-world synthetic biology experience

To Do

Test and optimize computationally designed biotin binding molecule.
Demonstrate selective secretion of proteins harboring secretion tag

Target Vector

[http://partsregistry.org/wiki/index.php/Part:BBa_K215002 BBa_K215002] Is a BioBrick part that was made to work in conjunction with the Secretion System ([http://partsregistry.org/wiki/index.php/Part:BBa_K215107 BBa_K215107]). When used it creates a fusion protein, that contains the prtB secretion signal recognized by the secretion system. It also fuses to the protein a nano-tag that is able to bind Streptavidin, TEV protease site (for isolating your protein from the fusion) and Histidine Epitote Tags for easy protein purification.

This part is composed of:
- [http://partsregistry.org/wiki/index.php/Part:BBa_K215000 BBa_K215000]: Protein Expression Vector
- [http://partsregistry.org/wiki/index.php/Part:BBa_K215001 BBa_K215001]: Fusion protein sequence

This part is used in:
- [http://partsregistry.org/wiki/index.php/Part:BBa_K215010 BBa_K215010]: GFP fusion protein
- [http://partsregistry.org/wiki/index.php/Part:BBa_K215011 BBa_K215011]: [http://partsregistry.org/wiki/index.php/Part:BBa_K215090 BBa_K215090] (organic phosphotriesterase) fusion protein

Current Status: Complete

Future directions:
- Make more universal, to work with all proteins
- Optimize secretion-tag and nano-tag for fusion proteins

Secretion System

[http://partsregistry.org/wiki/index.php/Part:BBa_K215107 BBa_K215107] Is a BioBrick version of the Type 1 secretion system of Erwinia Chrysanthemi. It secretes proteins that contain the prtB C-terminus Epitote tag. To create a fusion protein with the prtB tag see [http://partsregistry.org/wiki/index.php/Part:BBa_K215002 BBa_K215002].

This part is composed of
- [http://partsregistry.org/wiki/index.php/Part:BBa_K215100 BBa_K215100]: Gene encoding prtD
- [http://partsregistry.org/wiki/index.php/Part:BBa_K215101 BBa_K215101]: Gene encoding prtE
- [http://partsregistry.org/wiki/index.php/Part:BBa_K215102 BBa_K215102]: Gene encoding prtF
- [http://partsregistry.org/wiki/index.php/Part:BBa_K215103 BBa_K215103]: [http://partsregistry.org/wiki/index.php/Part:BBa_K21500 BBa_K21500] + [http://partsregistry.org/wiki/index.php/Part:BBa_K21501 BBa_K21501] prtD + prtE
- [http://partsregistry.org/wiki/index.php/Part:BBa_K215104 BBa_K215104]: [http://partsregistry.org/wiki/index.php/Part:BBa_K25103 BBa_K25103] + [http://partsregistry.org/wiki/index.php/Part:BBa_K25102 BBa_K25102] prtD + prtE + prtF
- [http://partsregistry.org/wiki/index.php/Part:BBa_K215105 BBa_K215105]: [http://partsregistry.org/wiki/index.php/Part:BBa_K215104 BBa_K215104] + [http://partsregistry.org/wiki/index.php/Part:BBa_B0014 BBa_B0014] in PSB1AK3 prtDEF + Terminator
- [http://partsregistry.org/wiki/index.php/Part:BBa_K215106 BBa_K215106]: [http://partsregistry.org/wiki/index.php/Part:BBa_K215104 BBa_K215104] + [http://partsregistry.org/wiki/index.php/Part:BBa_B0014 BBa_B0014] in PSB3T5 prtDEF + Terminator
- [http://partsregistry.org/wiki/index.php/Part:BBa_K215108 BBa_K215108]: [http://partsregistry.org/wiki/index.php/Part:BBa_J23114 BBa_J23114] + [http://partsregistry.org/wiki/index.php/Part:BBa_K215106 BBa_K215106] (Medium Promoter + prtDEF-Terminator)
- [http://partsregistry.org/wiki/index.php/Part:BBa_K215109 BBa_K215109]: [http://partsregistry.org/wiki/index.php/Part:BBa_J23113 BBa_J23113] + [http://partsregistry.org/wiki/index.php/Part:BBa_K215106 BBa_K215106] (Weak Promoter + prtDEF-Terminator)

Current Status: In progress

Future directions:
- Optimize RBS's
- Move secretion sequence into new plasmid, not TetR
- Try variety of cell lines.

Display System

[http://partsregistry.org/wiki/index.php/Part:BBa_K215200 BBa_K215200] is a Display vector that will allow for the presentation of any protein onto the surface of a cell by fusing the protein to Outer Membrane Protein A (OMPA). The fusion protein contains a GS peptide linker to space the displayed protein away from the outer membrane and also contains a TEV protease site to allow for purification of the displayed protein.

This Part is composed of:
- [http://partsregistry.org/wiki/index.php/Part:BBa_K215201 BBa_K215201]: Display Vector with 5 trans-membrane OMPA
- [http://partsregistry.org/wiki/index.php/Part:BBa_K215250 BBa_K215250]: Coding Sequence for fusion protein scaffold, 1 trans-membrane OMPA
- [http://partsregistry.org/wiki/index.php/Part:BBa_K215251 BBa_K215251]: Coding Sequence for fusion protein scaffold, 5 trans-membrane OMPA

This Part is used in:
- [http://partsregistry.org/wiki/index.php/Part:BBa_K215210 BBa_K215210]: Display system (1 trans-membrane) in protein expression vector
- [http://partsregistry.org/wiki/index.php/Part:BBa_K215211 BBa_K215211]: Display system (5 trans-membrane) in protein expression vector
- [http://partsregistry.org/wiki/index.php/Part:BBa_K215260 BBa_K215260]: [http://partsregistry.org/wiki/index.php/Part:BBa_K215210 BBa_K215210] fused to Streptavidin
- [http://partsregistry.org/wiki/index.php/Part:BBa_K215261 BBa_K215261]: [http://partsregistry.org/wiki/index.php/Part:BBa_K215211 BBa_K215211] fused to Streptavidin

Current Status: In progress

Future Directions:
- Use new CDS display vector to display streptavidin
- Design monomeric protein to bind peptide sequence
- Utilize other proteins on CDS to test for activity of other proteins

Biobricks Submitted	Current Status Report	Future Directions

Quick overview of what was done and the status of everyting. Maybe 3 paragraphs (display, secretion, conclusions)

@@ Line 7: / Line 7: @@
 ''Accomplished''
 *Create Protein Expression Vector for IPP system
-**<partinfo>BBa_K215000</partinfo>, Strong laq promoter with strong RBS
+**[http://partsregistry.org/wiki/index.php/Part:BBa_K215000 BBa_K215000], Strong laq promoter with strong RBS
 *Demonstrate Stable, Functional, His-Tagged Fusion protein from expression vector
 **[https://2009.igem.org/Team:Washington/Project/Target]See Project Page: Target
 *Synthesize and BioBrick type I secretion system from Erwinia Chrysanthemi
-**<partinfo>BBa_K215107</partinfo>, prtDEF from Erwinia Chrysanthemi under constitutive expression in low copy plasmid
+**[http://partsregistry.org/wiki/index.php/Part:BBa_K215107 BBa_K215107], prtDEF from Erwinia Chrysanthemi under constitutive expression in low copy plasmid
 *Develop assay for looking for secreted proteins
 **[https://2009.igem.org/Team:Washington/Project/Secretion]See Project Page: Secretion
 *Extensively characterize Legacy Surface Display BioBricks in registry[https://2009.igem.org/Team:Washington/Project/]
-**<partinfo>BBa_J36848</partinfo>
+**[http://partsregistry.org/wiki/index.php/Part:BBa_J36848 BBa_J36848]
-**<partinfo>BBa_J36849</partinfo>
+**[http://partsregistry.org/wiki/index.php/Part:BBa_J36849 BBa_J36849]
-**<partinfo>BBa_J36850</partinfo>
+**[http://partsregistry.org/wiki/index.php/Part:BBa_J36850 BBa_J36850]
-**<partinfo>BBa_J36851</partinfo>
+**[http://partsregistry.org/wiki/index.php/Part:BBa_J36851 BBa_J36851]
 *Develop new modular surface display system
 *Computationally design novel Biotin-binding protein using Rosetta
@@ Line 35: / Line 35: @@
 [[image:Target_structure.png |300px| left]]
-<partinfo>BBa_K215002</partinfo> Is a BioBrick part that was made to work in conjunction with the Secretion System (<partinfo>BBa_K215107</partinfo>). When used it creates a fusion protein, that contains the prtB secretion signal recognized by the secretion system. It also fuses to the protein a nano-tag that is able to bind Streptavidin, TEV protease site (for isolating your protein from the fusion) and Histidine Epitote Tags for easy protein purification.
+[http://partsregistry.org/wiki/index.php/Part:BBa_K215002 BBa_K215002] Is a BioBrick part that was made to work in conjunction with the Secretion System ([http://partsregistry.org/wiki/index.php/Part:BBa_K215107 BBa_K215107]). When used it creates a fusion protein, that contains the prtB secretion signal recognized by the secretion system. It also fuses to the protein a nano-tag that is able to bind Streptavidin, TEV protease site (for isolating your protein from the fusion) and Histidine Epitote Tags for easy protein purification.
 <br clear="all" />
 *This part is composed of:
-**<partinfo>BBa_K215000</partinfo>: Protein Expression Vector
+**[http://partsregistry.org/wiki/index.php/Part:BBa_K215000 BBa_K215000]: Protein Expression Vector
-**<partinfo>BBa_K215001</partinfo>: Fusion protein sequence
+**[http://partsregistry.org/wiki/index.php/Part:BBa_K215001 BBa_K215001]: Fusion protein sequence
 *This part is used in:
-**<partinfo>BBa_K215010</partinfo>: GFP fusion protein
+**[http://partsregistry.org/wiki/index.php/Part:BBa_K215010 BBa_K215010]: GFP fusion protein
-**<partinfo>BBa_K215011</partinfo>: <partinfo>BBa_K215090</partinfo> (organic phosphotriesterase) fusion protein
+**[http://partsregistry.org/wiki/index.php/Part:BBa_K215011 BBa_K215011]: [http://partsregistry.org/wiki/index.php/Part:BBa_K215090 BBa_K215090] (organic phosphotriesterase) fusion protein
 *Current Status: Complete
@@ Line 57: / Line 57: @@
 [[image:Secretion_structure.png |300px | left]]
-<partinfo>BBa_K215107</partinfo> Is a BioBrick version of the Type 1 secretion system of Erwinia Chrysanthemi. It secretes proteins that contain the prtB C-terminus Epitote tag. To create a fusion protein with the prtB tag see <partinfo>BBa_K215002</partinfo>.
+[http://partsregistry.org/wiki/index.php/Part:BBa_K215107 BBa_K215107] Is a BioBrick version of the Type 1 secretion system of Erwinia Chrysanthemi. It secretes proteins that contain the prtB C-terminus Epitote tag. To create a fusion protein with the prtB tag see [http://partsregistry.org/wiki/index.php/Part:BBa_K215002 BBa_K215002].
 <br clear="all" />
 *This part is composed of
-**<partinfo>BBa_K215100</partinfo>: Gene encoding prtD
+**[http://partsregistry.org/wiki/index.php/Part:BBa_K215100 BBa_K215100]: Gene encoding prtD
-**<partinfo>BBa_K215101</partinfo>: Gene encoding prtE
+**[http://partsregistry.org/wiki/index.php/Part:BBa_K215101 BBa_K215101]: Gene encoding prtE
-**<partinfo>BBa_K215102</partinfo>: Gene encoding prtF
+**[http://partsregistry.org/wiki/index.php/Part:BBa_K215102 BBa_K215102]: Gene encoding prtF
-**<partinfo>BBa_K215103</partinfo>: <partinfo>BBa_K21500</partinfo> + <partinfo>BBa_K21501</partinfo> prtD + prtE
+**[http://partsregistry.org/wiki/index.php/Part:BBa_K215103 BBa_K215103]: [http://partsregistry.org/wiki/index.php/Part:BBa_K21500 BBa_K21500] + [http://partsregistry.org/wiki/index.php/Part:BBa_K21501 BBa_K21501] prtD + prtE
-**<partinfo>BBa_K215104</partinfo>: <partinfo>BBa_K25103</partinfo> + <partinfo>BBa_K25102</partinfo> prtD + prtE + prtF
+**[http://partsregistry.org/wiki/index.php/Part:BBa_K215104 BBa_K215104]: [http://partsregistry.org/wiki/index.php/Part:BBa_K25103 BBa_K25103] + [http://partsregistry.org/wiki/index.php/Part:BBa_K25102 BBa_K25102] prtD + prtE + prtF
-**<partinfo>BBa_K215105</partinfo>: <partinfo>BBa_K215104</partinfo> + <partinfo>BBa_B0014</partinfo> in PSB1AK3 prtDEF + Terminator
+**[http://partsregistry.org/wiki/index.php/Part:BBa_K215105 BBa_K215105]: [http://partsregistry.org/wiki/index.php/Part:BBa_K215104 BBa_K215104] + [http://partsregistry.org/wiki/index.php/Part:BBa_B0014 BBa_B0014] in PSB1AK3 prtDEF + Terminator
-**<partinfo>BBa_K215106</partinfo>: <partinfo>BBa_K215104</partinfo> + <partinfo>BBa_B0014</partinfo> in PSB3T5 prtDEF + Terminator
+**[http://partsregistry.org/wiki/index.php/Part:BBa_K215106 BBa_K215106]: [http://partsregistry.org/wiki/index.php/Part:BBa_K215104 BBa_K215104] + [http://partsregistry.org/wiki/index.php/Part:BBa_B0014 BBa_B0014] in PSB3T5 prtDEF + Terminator
-**<partinfo>BBa_K215108</partinfo>: <partinfo>BBa_J23114</partinfo> + <partinfo>BBa_K215106</partinfo> (Medium Promoter + prtDEF-Terminator)
+**[http://partsregistry.org/wiki/index.php/Part:BBa_K215108 BBa_K215108]: [http://partsregistry.org/wiki/index.php/Part:BBa_J23114 BBa_J23114] + [http://partsregistry.org/wiki/index.php/Part:BBa_K215106 BBa_K215106] (Medium Promoter + prtDEF-Terminator)
-**<partinfo>BBa_K215109</partinfo>: <partinfo>BBa_J23113</partinfo> + <partinfo>BBa_K215106</partinfo> (Weak Promoter + prtDEF-Terminator)
+**[http://partsregistry.org/wiki/index.php/Part:BBa_K215109 BBa_K215109]: [http://partsregistry.org/wiki/index.php/Part:BBa_J23113 BBa_J23113] + [http://partsregistry.org/wiki/index.php/Part:BBa_K215106 BBa_K215106] (Weak Promoter + prtDEF-Terminator)
 *Current Status: In progress
@@ Line 83: / Line 83: @@
 [[image:Display_structure.png |300px | left]]
-<partinfo>BBa_K215200</partinfo> is a Display vector that will allow for the presentation of any protein onto the surface of a cell by fusing the protein to Outer Membrane Protein A (OMPA). The fusion protein contains a GS peptide linker to space the displayed protein away from the outer membrane and also contains a TEV protease site to allow for purification of the displayed protein.
+[http://partsregistry.org/wiki/index.php/Part:BBa_K215200 BBa_K215200] is a Display vector that will allow for the presentation of any protein onto the surface of a cell by fusing the protein to Outer Membrane Protein A (OMPA). The fusion protein contains a GS peptide linker to space the displayed protein away from the outer membrane and also contains a TEV protease site to allow for purification of the displayed protein.
 <br clear="all" />
 *This Part is composed of:
-**<partinfo>BBa_K215201</partinfo>: Display Vector with 5 trans-membrane OMPA
+**[http://partsregistry.org/wiki/index.php/Part:BBa_K215201 BBa_K215201]: Display Vector with 5 trans-membrane OMPA
-**<partinfo>BBa_K215250</partinfo>: Coding Sequence for fusion protein scaffold, 1 trans-membrane OMPA
+**[http://partsregistry.org/wiki/index.php/Part:BBa_K215250 BBa_K215250]: Coding Sequence for fusion protein scaffold, 1 trans-membrane OMPA
-**<partinfo>BBa_K215251</partinfo>: Coding Sequence for fusion protein scaffold, 5 trans-membrane OMPA
+**[http://partsregistry.org/wiki/index.php/Part:BBa_K215251 BBa_K215251]: Coding Sequence for fusion protein scaffold, 5 trans-membrane OMPA
 *This Part is used in:
-**<partinfo>BBa_K215210</partinfo>: Display system (1 trans-membrane) in protein expression vector
+**[http://partsregistry.org/wiki/index.php/Part:BBa_K215210 BBa_K215210]: Display system (1 trans-membrane) in protein expression vector
-**<partinfo>BBa_K215211</partinfo>: Display system (5 trans-membrane) in protein expression vector
+**[http://partsregistry.org/wiki/index.php/Part:BBa_K215211 BBa_K215211]: Display system (5 trans-membrane) in protein expression vector
-**<partinfo>BBa_K215260</partinfo>: <partinfo>BBa_K215210</partinfo> fused to Streptavidin
+**[http://partsregistry.org/wiki/index.php/Part:BBa_K215260 BBa_K215260]: [http://partsregistry.org/wiki/index.php/Part:BBa_K215210 BBa_K215210] fused to Streptavidin
-**<partinfo>BBa_K215261</partinfo>: <partinfo>BBa_K215211</partinfo> fused to Streptavidin
+**[http://partsregistry.org/wiki/index.php/Part:BBa_K215261 BBa_K215261]: [http://partsregistry.org/wiki/index.php/Part:BBa_K215211 BBa_K215211] fused to Streptavidin
 *Current Status: In progress