Team:Edinburgh/Project

From 2011.igem.org

(Difference between revisions)

Latest revision as of 14:41, 28 July 2011

This page is obsolete. The project should be documented at the relevant subpages:

Designs

The general formats for the basic phage and cell display would be:

Promoter-RBS-LeaderSequence-Enzyme-(Linker?)-pVIII
Promoter-RBS-INP-(Linker?)-Enzyme

Actions that ought to be carried out

General

Make or acquire fusion-ready cellulases (e.g. <partinfo>BBa_K392006</partinfo>, <partinfo>BBa_K392007</partinfo>, <partinfo>BBa_K392008</partinfo>).

Since a simple test system is desired, make fusion-ready amylase, from E. coli's own malS gene.

Make or acquire spacer/linker sequences?
- Short linkers could be ordered as oligos. Berkeley 2009 made some longer linkers which we could order.

Phage display

Acquire M13 phage.

PCR from our [http://www.neb.com/nebecomm/products/productn4018.asp M13mp18] to get:
- pVIII mature protein
- pVIII leader sequence, coding for: MKKSLVLKASVAVATLVPMLSFA.

Make or acquire fusion-ready pIII gene? (optional)
- See <partinfo>BBa_K415138</partinfo> (complete) or <partinfo>BBa_K257001</partinfo> (sans leader sequence).
- Our final fusion will need the leader sequence followed by the gene of interest followed by pIII proper.

Cell display

Make or acquire fusion-ready cell-surface display parts.
- See Berkeley 2009 (but perhaps ignore them).
- Instead, <partinfo>BBa_K265008</partinfo> (ice nucleation protein) looks promising.
- The fliC gene seems to be the target for flagella display; note that the displayed protein has to be inserted into the middle of it.

Biorefinery

To add...

Completion

Fuse whatever enzymes to cell-display parts or pVIII or maybe pIII.

???

Profit!

@@ Line 1: / Line 1: @@
-A [[Project descriptions|project description]] and [[Safety/Proposals|safety proposal]] is supposed to be submitted by July 15.
+{{:Team:Edinburgh/tech/Navbox}}
+<html><script type="text/javascript" >$(document).ready(function() {
+	getMenus('home', 'home_project');
+}); </script></html>
+<div class="main_body">
+__NOTOC__
+'''This page is obsolete. The project should be documented at the relevant subpages:'''
-==Project abstract==
+* [[Team:Edinburgh/Phage Reactors 2.0|Phage Reactors]]
+* [[Team:Edinburgh/Cell Display|Cell Display]]
+* [[Team:Edinburgh/Biorefineries|Biorefineries]]
-This year we will create "reactors", consisting of scaffolds for various enzymes, to carry out reactions in an extracellular environment. The hope is that, by combining the activity of multiple synergistic enzymes in a small space, high efficiency will be achieved. At least two different systems will be investigated:
+==Designs==
-* Using bacteria as the scaffold, and attaching enzymes by cell-surface display techniques.
+The general formats for the basic phage and cell display would be:
-* Using M13 phage as the scaffold, and attaching enzymes by cell-surface display techniques to the pVIII coat protein.
-** Additionally, it may be possible to attach multiple phage to beads, making a larger "reactor".
-As an example system, we will try creating ''E. coli'' with different cellulase enzymes displayed on the surface, as well as (if possible) M13 phage with the same enzymes displayed on the pVIII coat protein.
+* Promoter-RBS-LeaderSequence-Enzyme-(Linker?)-pVIII
+* Promoter-RBS-INP-(Linker?)-Enzyme
-==Pages==
+==Actions that ought to be carried out==
-* [[Team:Edinburgh/Phage Reactors 2.0|Phage Reactors]]
+===General===
-* [[Team:Edinburgh/Cell Display|Cell Display]]
-==To do==
 * Make or acquire fusion-ready cellulases (e.g. <partinfo>BBa_K392006</partinfo>, <partinfo>BBa_K392007</partinfo>, <partinfo>BBa_K392008</partinfo>).
+* Since a simple test system is desired, make fusion-ready amylase, from ''E. coli's'' own ''malS'' gene.
+* Make or acquire spacer/linker sequences?
+** Short linkers could be ordered as oligos. [[Team:Edinburgh/Berkeley 2009 Parts|Berkeley 2009 made some longer linkers]] which we could order.
+===Phage display===
 * Acquire M13 phage.
-* If a simple test system is desired, make fusion-ready amylase?
+* PCR from our [http://www.neb.com/nebecomm/products/productn4018.asp M13mp18] to get:
+** pVIII mature protein
+** pVIII leader sequence, coding for: MKKSLVLKASVAVATLVPMLSFA.
-* Make or acquire fusion-ready pVIII gene (perhaps from <partinfo>BBa_K415151</partinfo>).
+* Make or acquire fusion-ready pIII gene? (optional)
-** Remember it has a leader sequence. Our final fusion will need the leader sequence followed by the gene of interest followed by pVIII proper.
+** See <partinfo>BBa_K415138</partinfo> (complete) or <partinfo>BBa_K257001</partinfo> (''sans'' leader sequence).
+** Our final fusion will need the leader sequence followed by the gene of interest followed by pIII proper.
-* Make or acquire fusion-ready pIII gene?
+===Cell display===
-** Remember it has a leader sequence. Our final fusion will need the leader sequence followed by the gene of interest followed by pIII proper.
-* Make or acquire fusion-ready cell-surface display parts (see [[Team:Berkeley Wetlab/Cell Surface Display Parts|Berkeley 2009]]).
+* Make or acquire fusion-ready cell-surface display parts.
+** See [https://2009.igem.org/Team:Berkeley_Wetlab/Cell_Surface_Display_Parts Berkeley 2009] (but perhaps ignore them).
+** Instead, <partinfo>BBa_K265008</partinfo> (ice nucleation protein) looks promising.
+** The ''fliC'' gene seems to be the target for flagella display; note that the displayed protein has to be inserted into the middle of it.
+===Biorefinery===
+:''To add...''
+===Completion===
 * Fuse whatever enzymes to cell-display parts or pVIII or maybe pIII.
+* ???
+* '''Profit!'''
-{{:Team:Edinburgh/Template:Navbox}}
+</div> <!-- /main_body-->
+<html></div> <!-- /mids --></html>