Difference between revisions of "Team:Vilnius-Lithuania/Design"

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                     <strong>Fig 1</strong> The composition of a liposome with encapsulated machinery for membrane protein  
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                    integration. Size, membrane composition and interior composition can be easily varied.
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                                      <img src="https://static.igem.org/mediawiki/2018/3/3b/T--Vilnius-Lithuania--Fig1_Liposomes.png"/>
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                                      <strong>Fig 1</strong> he composition of a liposome with encapsulated machinery for membrane protein integration. Size, membrane composition and interior composition can be easily varied.
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                     The first step we had to take towards the production of liposomes was to design, fabricate and prepare a unique microfluidic device. A microfluidic channel design was created with the AutoCAD platform. The prototype acts as a photomask during the photolithography to create a master for the fabrication of microfluidic chips.  Our design consists of an array of 16 separate microfluidic channel devices distributed parallelly in groups of four on a single chip (Fig. 2). The dimensions of the microchannels limit the size range of the synthesized liposomes. To dig deeper into the details of how the dimensions of the microfluidic channels influence liposome size we created a phase-field based <strong><var><a href="https://2018.igem.org/Team:Vilnius-Lithuania/Model#COMSOL_model">SynFlow</a></var></strong> model with COMSOL Multiphysics. Auxiliary parametric sweeps were performed that defined the dimensions needed to attain cell – sized liposomes, a range from 5 µm to 30 µm. The final design can be downloaded here and used by anyone interested in the synthesis of liposomes.
 
                     The first step we had to take towards the production of liposomes was to design, fabricate and prepare a unique microfluidic device. A microfluidic channel design was created with the AutoCAD platform. The prototype acts as a photomask during the photolithography to create a master for the fabrication of microfluidic chips.  Our design consists of an array of 16 separate microfluidic channel devices distributed parallelly in groups of four on a single chip (Fig. 2). The dimensions of the microchannels limit the size range of the synthesized liposomes. To dig deeper into the details of how the dimensions of the microfluidic channels influence liposome size we created a phase-field based <strong><var><a href="https://2018.igem.org/Team:Vilnius-Lithuania/Model#COMSOL_model">SynFlow</a></var></strong> model with COMSOL Multiphysics. Auxiliary parametric sweeps were performed that defined the dimensions needed to attain cell – sized liposomes, a range from 5 µm to 30 µm. The final design can be downloaded here and used by anyone interested in the synthesis of liposomes.
 
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             <p><strong>Fig. 2 a</strong> AutoCAD design for the photomask. There are 16 individual microchannel devices on a
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                    single chip. <strong>b</strong> One device consists of three inlets, an outlet and a star-shaped junction.</p>
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                                      <img src="https://static.igem.org/mediawiki/2018/3/3b/T--Vilnius-Lithuania--Fig1_Liposomes.png"/>
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                                      <strong>Fig 2 a</strong>AutoCAD design for the photomask. There are 16 individual microchannel devices on a
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                                      single chip. <strong>b</strong> One device consists of three inlets, an outlet and a star-shaped junction.
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                     <h2>Photolithography as a tool for microfluidic chip fabrication</h2>
 
                     <h2>Photolithography as a tool for microfluidic chip fabrication</h2>
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                     <li>Chikh, G., Li, W., Schutze-Redelmeier, M., Meunier, J. & Bally, M. Attaching histidine-tagged peptides and proteins to lipid-based carriers through use of metal-ion-chelating lipids. Biochimica et Biophysica Acta (BBA) - Biomembranes 1567, 204-212 (2002).
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                     <li>1.Chikh, G., Li, W., Schutze-Redelmeier, M., Meunier, J. & Bally, M. Attaching histidine-tagged peptides and proteins to lipid-based carriers through use of metal-ion-chelating lipids. Biochimica et Biophysica Acta (BBA) - Biomembranes 1567, 204-212 (2002).
 
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                     <li>Blanchette, C., Fischer, N., Corzett, M., Bench, G. & Hoeprich, P. Kinetic Analysis of His-Tagged Protein Binding to Nickel-Chelating Nanolipoprotein Particles. Bioconjugate Chemistry 21, 1321-1330 (2010).
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                     <li>2.Blanchette, C., Fischer, N., Corzett, M., Bench, G. & Hoeprich, P. Kinetic Analysis of His-Tagged Protein Binding to Nickel-Chelating Nanolipoprotein Particles. Bioconjugate Chemistry 21, 1321-1330 (2010).
 
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                     <li>Ederth, J., Mandava, C., Dasgupta, S. & Sanyal, S. A single-step method for purification of active His-tagged ribosomes from a genetically engineered Escherichia coli. Nucleic Acids Research 37, e15-e15 (2008).
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                     <li>3.Ederth, J., Mandava, C., Dasgupta, S. & Sanyal, S. A single-step method for purification of active His-tagged ribosomes from a genetically engineered Escherichia coli. Nucleic Acids Research 37, e15-e15 (2008).
 
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                     <li>Spillmann, S. & Nierhaus, K. The ribosomal protein L24 of Escherichia coli is an assembly protein. Journal of Biological Chemistry (1978). at http://www.jbc.org/content/253/19/7047.long     
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                     <li>4.Spillmann, S. & Nierhaus, K. The ribosomal protein L24 of Escherichia coli is an assembly protein. Journal of Biological Chemistry (1978). at http://www.jbc.org/content/253/19/7047.long     
 
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                     <li>GU, S. The signal recognition particle binds to protein L23 at the peptide exit of the Escherichia coli ribosome. RNA 9, 566-573 (2003).
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                     <li>5.GU, S. The signal recognition particle binds to protein L23 at the peptide exit of the Escherichia coli ribosome. RNA 9, 566-573 (2003).
 
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                     <li>STOFFLER-MEILICKE, M., DABBS, E., ALBRECHT-EHRLICH, R. & STOFFLER, G. A mutant from Escherichia coli which lacks ribosomal proteins S17 and L29 used to localize these two proteins on the ribosomal surface. European Journal of Biochemistry 150, 485-490 (1985).
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                     <li>6.STOFFLER-MEILICKE, M., DABBS, E., ALBRECHT-EHRLICH, R. & STOFFLER, G. A mutant from Escherichia coli which lacks ribosomal proteins S17 and L29 used to localize these two proteins on the ribosomal surface. European Journal of Biochemistry 150, 485-490 (1985).
 
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                     <li>Noeske, J. et al. Synergy of Streptogramin Antibiotics Occurs Independently of Their Effects on Translation. Antimicrobial Agents and Chemotherapy 58, 5269-5279 (2014).
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                     <li>7.Noeske, J. et al. Synergy of Streptogramin Antibiotics Occurs Independently of Their Effects on Translation. Antimicrobial Agents and Chemotherapy 58, 5269-5279 (2014).
 
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                     <li>Jiang, Y. et al. Multigene Editing in the Escherichia coli Genome via the CRISPR-Cas9 System. Applied and Environmental Microbiology 81, 2506-2514 (2015)
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                     <li>8.Jiang, Y. et al. Multigene Editing in the Escherichia coli Genome via the CRISPR-Cas9 System. Applied and Environmental Microbiology 81, 2506-2514 (2015)
 
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                 </ol>
 
                 </ol>

Revision as of 00:04, 18 October 2018

Design and Results

Results

Cell-free, synthetic biology systems open new horizons in engineering biomolecular systems which feature complex, cell-like behaviors in the absence of living entities. Having no superior genetic control, user-controllable mechanisms to regulate gene expression are necessary to successfully operate these systems. We have created a small collection of synthetic RNA thermometers that enable temperature-dependent translation of membrane proteins, work well in cells and display great potential to be transferred to any in vitro protein synthesis system.

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