Difference between revisions of "Team:UC San Diego/Parts"

 
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                   <h1>List of Parts</h1>
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The collection of parts described herein forms a modular platform to determine which of the enzymes of the fatty acid reductase complex is rate limiting. Each permutation of the complex’s genes is compatible for coexpression with the <i>lux</i>AB construct. This methodology of stoichiometric expression can also be used to determine rate-limiting steps in other metabolic systems.
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The part codes for the fatty acid reductase complex of the bacterial lux system from Photobacterium Phosphoreum. The system recruits fatty acids and converts them into aldehyde to be used as substrate in the system's light producing reaction. The genes coding for each subunit of the complex are separated by a P2A sequence for stoichiometric expression. The part is comprised of three unique genes - LuxC, LuxD, and LuxE, coding for a reductase, transferase, and synthetase, respectively. Each gene has an attached antibody tag - LuxC has a FLAG tag, LuxD has a V5 tag, and LuxE has a HA tag. A duplicate copy of LuxC is present, resulting in its overexpression. Genes used are codon optimized for expression in yeast.<br><br>
 
  
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Each of the genes used was derived from the marine bioluminescent bacterium Photobacterium Phosphoreum. <br><br>
 
  
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The coding sequence was too large to be synthesized in one piece, so it was instead synthesized in a series of fragments that were assembled via Gibson Assembly. </p>
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                   <h1>K1842000</h1>
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                   <h3>K1842000</h3>
 
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<p>LuxABfrp - Coding - Engineered LuxAB with Vibrio Harveyi frp; 2A linker present between each protein for stoichiometric expression.  
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<p><b>LuxABfrp - Coding</b> - Engineered <i>lux</i>AB with <i>Vibrio Harveyi frp</i>; 2A linker present between each protein for stoichiometric expression.  
 
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LuxCDEC - Coding - Photobacterium Phosphoreum fatty acid reductase complex with overexpressed synthetase; 2A linker present between each protein for stoichiometric expression. </p>
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<b>LuxCDEC - Coding</b> - <i>Photobacterium Phosphoreum</i> fatty acid reductase complex with overexpressed reductase; 2A linker present between each protein for stoichiometric expression. </p>
  
  
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LuxCDED - Coding -  Photobacterium Phosphoreum fatty acid reductase complex with overexpressed transferase; 2A linker present between each protein for stoichiometric expression.  
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<b>LuxCDED - Coding</b> -  <i>Photobacterium Phosphoreum</i> fatty acid reductase complex with overexpressed transferase; 2A linker present between each protein for stoichiometric expression.  
 
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<p>LuxCDEE - Coding - Photobacterium Phosphoreum fatty acid reductase complex with overexpressed reductase; 2A linker present between each protein for stoichiometric expression.  
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<p><b>LuxCDEE - Coding</b> - <i>Photobacterium Phosphoreum</i> fatty acid reductase complex with overexpressed synthetase; 2A linker present between each protein for stoichiometric expression.  
 
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                  <h3>K1842004</h3>
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<p><b>LuxDEC - Coding</b> - <i>Photobacterium Phosphoreum</i> fatty acid reductase complex with even stoichiometry for LuxCDE. 2A linker present between each protein for stoichiometric expression.
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Latest revision as of 21:19, 20 November 2015


List of Parts

The collection of parts described herein forms a modular platform to determine which of the enzymes of the fatty acid reductase complex is rate limiting. Each permutation of the complex’s genes is compatible for coexpression with the luxAB construct. This methodology of stoichiometric expression can also be used to determine rate-limiting steps in other metabolic systems.

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K1842000

LuxABfrp - Coding - Engineered luxAB with Vibrio Harveyi frp; 2A linker present between each protein for stoichiometric expression.

K1842001

LuxCDEC - Coding - Photobacterium Phosphoreum fatty acid reductase complex with overexpressed reductase; 2A linker present between each protein for stoichiometric expression.

K1842002

LuxCDED - Coding - Photobacterium Phosphoreum fatty acid reductase complex with overexpressed transferase; 2A linker present between each protein for stoichiometric expression.

K1842003

LuxCDEE - Coding - Photobacterium Phosphoreum fatty acid reductase complex with overexpressed synthetase; 2A linker present between each protein for stoichiometric expression.

K1842004

LuxDEC - Coding - Photobacterium Phosphoreum fatty acid reductase complex with even stoichiometry for LuxCDE. 2A linker present between each protein for stoichiometric expression.