Team:ETH Zurich/Project/Medal Relevant

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Latest revision as of 05:15, 30 October 2008

Medal Relevant Issues

Bronze Relevant

Register the team, have a great summer, and have fun attending the Jamboree

All registration tasks were done, as can be seen here and we hope to have fun at Jamboree.

Successfully complete and submit a Project Summary form

You can see our submitted Abstract here

Create and share a Description of the team's project via the iGEM wiki

The general project description can be found on this page of the wiki.

Present a Poster and Talk at the iGEM Jamboree

Enter information detailing at least one new standard BioBrick Part or Device in the Registry of Parts

We have deposited information detailing over [http://partsregistry.org/cgi/partsdb/pgroup.cgi?pgroup=iGEM2008&group=ETH_Zurich 40 novel] BioBricks in the registry. Among them T4 Ligase, SceI, LacI IS repressor mutants, RMF are new to the registry.

Submit DNA for at least one new BioBrick Part or Device to the Registry of Parts

Shipped via DHL on the 29th of October. Tracking Number: 4531864046

Silver Relevant

Demonstrate that at least one new BioBrick Part or Device of your own design and construction works as expected

LacI IS mutants

Based on atomic structures of LacI and evidence from genetic studies we have constructed a set of eight mutants of LacI, which are unresponsive to IPTG or lactose and are a useful component of genetic circuits or pulse generators. Qualitative genetic experiments were conducted to estimate the strength of repression exerted by the LacI IS mutants and the mutants were found to repress expression from lac-controlled promoters even at 10mM IPTG. We have provided the LacI IS mutants by themselves (BioBricks [http://partsregistry.org/wiki/index.php?title=Part:BBa_K142000 K142000] through [http://partsregistry.org/wiki/index.php?title=Part:BBa_K142007 K142007]) and with RBS and terminator (BioBricks [http://partsregistry.org/wiki/index.php?title=Part:BBa_K142008 K142008] through [http://partsregistry.org/wiki/index.php?title=Part:BBa_K142015 K142015]) for straigthforward integration into genetic circuits.

Ribosome Modulation Factor (RMF)

We have introduced RMF as a new BioBrick ([http://partsregistry.org/wiki/index.php?title=Part:BBa_K142040 BBa_K142040] and [http://partsregistry.org/wiki/index.php?title=Part:BBa_K142041 K142041]). Based on extensive literature search as well as our own biophysical experiments we concluded that RMF would among other uses be helpful as "pause switch", since it terminates protein expression rapidly and reversibly without killing the cell.

Characterize the operation of at least one new BioBrick Part or Device and enter this information on the Parts or Device page via the Registry of Parts

The LacI IS mutants and the devices constructed from them have been documented in the registry.

Gold Relevant

Analysis, modeling, and simulation of BioBrick Parts or Devices

We propose a novel method of random gene deletion and chemostat-based selection of species with a reduced genome. For this we provide an algorithm described here.

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 == Medal Relevant Issues ==
-=== Silver Relevant ===
+=== Bronze Relevant ===
+=====Register the team, have a great summer, and have fun attending the Jamboree=====
+All registration tasks were done, as can be seen [https://igem.org/Team.cgi?id=142 here] and we hope to have fun at Jamboree.
-=== Gold Relevant ===
+=====Successfully complete and submit a Project Summary form=====
+You can see our submitted Abstract [https://2008.igem.org/Team:ETH_Zurich#Project_Abstract here]
-==== Analysis, modeling, and simulation of BioBrick Parts or Devices ====
+=====Create and share a Description of the team's project via the iGEM wiki=====
-We propose a novel method of random gene deletion and chemostat-based selection of species with a reduced genome. For this we provide an algorithm described below.
+The general project description can be found on [https://2008.igem.org/Team:ETH_Zurich/Project/Overview this page] of the wiki.
-[[Image:Gold_Algorithm.jpg|frame|none|Genome Reduction Algorithm]]
+=====Present a Poster and Talk at the iGEM Jamboree=====
-===== Modeling Framework =====
+=====Enter information detailing at least one new standard BioBrick Part or Device in the Registry of Parts=====
-This algorithm requires a modeling framework consisting of four main parts:
+We have deposited information detailing over [http://partsregistry.org/cgi/partsdb/pgroup.cgi?pgroup=iGEM2008&group=ETH_Zurich 40 novel] BioBricks in the registry. Among them T4 Ligase, SceI, LacI IS repressor mutants, RMF are new to the registry.
-# [[Team:ETH_Zurich/Modeling/Genome_Static_Analysis|Statistical Analysis]] of DNA fragments produced by Restriction Enzymes
-# [[Team:ETH_Zurich/Modeling/Genome-Scale_Model| Flux Balance Analysis]] on a Genome Scale Model
-# [[Team:ETH_Zurich/Modeling/Chemostat_Selection|Growth Simulations in Chemostat]]
-# [[Team:ETH_Zurich/Modeling/Switch_Circuit|Switch generator]] for short-time Restriction Enzyme Expression
-===== Detailed description of the Modeling Framework =====
+=====Submit DNA for at least one new BioBrick Part or Device to the Registry of Parts=====
-# '''Restriction Enzyme Analysis'''
+Shipped via DHL on the 29th of October. Tracking Number: 4531864046
-#:We want to identify a restriction enzyme for genome reduction that maximizes the probability of genome reduction and minimizes the probability to hit an (known) essential gene.  Therefore, given a genome data of an E. Coli strain, the genome is digested using ca. 700 different restriction enzymes (with different recognition patterns). The resulting fragments are analyzed using the available annotation of the genome. The number of genes disrupted is calculated for each fragment. Statistical measures of the restriction enzyme effects, such as average size of the fragments and its variance, and the average number of (all and only essential) genes and its variance are calculated. According to these results, the most suitable restriction enzyme is chosen.
-# '''Genome Scale Model and Flux Balance Analysis'''
-#:We apply Flux Balance Analysis on the state-of-the-art genome scale model for E.Coli iAF1260 (1260 genes included) in order to calculate the biomass yield (and therefore the corresponding growth rate) for wildtype and reduced strains. In order to apply a selective condition for reduced genome strain, the model is modified by constraining nucleotide availability: this means inactivation of a part of a thymidine synthesis pathway and external uptake of thymidine from the medium. The optimal external thymidine concentration is determined which enables the fastest change in growth rate of a reduced genome mutant.
-# '''Growth Simulations in Chemostat'''
-#:We model a chemostat using a system of coupled differential equations for a population of different mutants with different growth rates competing for a limited external thymidine concentration. By varying the time intervals between two pulses of restriction enzymes (where only a small amount of previous population survived and new mutants are generated) we control the number of different mutants considering for further gene deletions after the next pulse event, consequently the diversity of a population.
-# '''Switch generator for short-time Restriction Enzyme Expression'''
-#:Using a novel pulsing mechanism consisting of two signals, start signal, which initiates the expression of restriction enzymes and stop signal, which switches off the expression, we are able to delete genome fragments in vivo. To simulate this system we developed a switch curcuit which works as follows (see [[Media:SimpleCircuit.jpg| Switching Curcuit]]) by inducing the system with IPTG, the restriction enzyme (RE), which is under control of lacI, can be expressed. In order to stop the expression the system is induced with tet, which inhibits the binding of tetR to LacIS (a mutant of LacI, which is not inducable by IPTG),  therefore activates the  LacIS expression and consecutive termination of RE expression. This curcuit is modeled by ca. 40 reactions and is simulated using ODE solver and stochastic simulations.
-===== Summary of the Algorithm and Interplay of Frameworks Componets =====
+=== Silver Relevant ===
+=====Demonstrate that at least one new BioBrick Part or Device of your own design and construction works as expected=====
-First tree steps initialize and prepare the system for gene deletions and growth simulations.
+======LacI IS mutants======
-In the first step the genome data are analyzed in order to find a most suitable restriction enzyme for random fragments deletion using the  “Restriction Enzyme Analysis”- procedure.  For the second step the state-of-the art model is adjusted by introducing a selective pressure due to the genome size. Thirdly, initial population consists of one type, namely wildtype.
+Based on atomic structures of LacI and evidence from genetic studies we have constructed a set of eight mutants of LacI, which are unresponsive to IPTG or lactose and are a useful component of genetic circuits or pulse generators. [[Team:ETH_Zurich/Wetlab/Switch_Circuit|Qualitative genetic experiments]] were conducted to estimate the strength of repression exerted by the LacI IS mutants and the mutants were found to repress expression from lac-controlled promoters even at 10mM IPTG. We have provided the LacI IS mutants by themselves (BioBricks [http://partsregistry.org/wiki/index.php?title=Part:BBa_K142000 K142000] through [http://partsregistry.org/wiki/index.php?title=Part:BBa_K142007 K142007]) and with RBS and terminator (BioBricks [http://partsregistry.org/wiki/index.php?title=Part:BBa_K142008 K142008] through [http://partsregistry.org/wiki/index.php?title=Part:BBa_K142015 K142015]) for straigthforward integration into genetic circuits.
-The next steps of the algorithm perform in depth genome fragments deletion and growth simulation.
+======Ribosome Modulation Factor (RMF)======
-First, we simulate the restriction enzyme expression and the consecutive population decline and occurrence of new mutants with reduced genome, which growth rates can be predicted using Flux Balance Analysis. For these simulations we used the framework parts: “Switch generator” and “Flux Balance Analysis on a Genome Scale Model “.
+We have introduced RMF as a new BioBrick ([http://partsregistry.org/wiki/index.php?title=Part:BBa_K142040 BBa_K142040] and [http://partsregistry.org/wiki/index.php?title=Part:BBa_K142041 K142041]). Based on extensive literature search as well as our own [[Team:ETH_Zurich/Wetlab/Switch_Circuit|biophysical experiments]] we concluded that RMF would among other uses be helpful as "pause switch", since it terminates protein expression rapidly and reversibly without killing the cell.
-Secondly, the growth simulations are performed using a chemostat model and the distribution of different mutant types after the growth type are obtained.
-This simulation continues until no better mutants can be generated.
-Eventually, the genome data of the fastest growing reduced genome mutant can be returned.
+=====Characterize the operation of at least one new BioBrick Part or Device and enter this information on the Parts or Device page via the Registry of Parts=====
+The LacI IS mutants and the devices constructed from them have been documented in the registry.
+=== Gold Relevant ===
+==== Analysis, modeling, and simulation of BioBrick Parts or Devices ====
+We propose a novel method of random gene deletion and chemostat-based selection of species with a reduced genome. For this we provide an algorithm described [https://2008.igem.org/Team:ETH_Zurich/Modeling/Framework here].
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