Difference between revisions of "Part:BBa K2533048"
 
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<partinfo>BBa_K2533048 short</partinfo>
 
<partinfo>BBa_K2533048 short</partinfo>
  
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Production of lactate and transport lactic acid
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malate dehydrogenase & L-lactate permease
 
<h1>'''Usage and biology'''</h1>
 
<h1>'''Usage and biology'''</h1>
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dld refers to FAD-dependent D-lactate dehydrogenase which could catalyze D-lactate’s transformation into pyruvate. With the overexpression of dld, Shewanella could utilize D-lactate more efficiently, which brings more electricity being produced.
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It encodes malate dehydrogenase and L-lactate permease, which could produce and transport lactate. mleS could convert malic acid to L-lactate. lldP could transport lactate out of the cell. In this way, Rhodopseudomonas palustris could efficiently transport produced lactate out of the cell, which later will be used by Shewanella.
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<h1>'''Characterization'''</h1>
 
<h1>'''Characterization'''</h1>
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This is one section for lactate utilization part.
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This is one section for lactate production and transportation part.
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[[File:T--HUST-China--2018-tonglu-mles-lldp.png ‎|400px|thumb|center|Figure1:RBS-mleS-RBS-lldP]]
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[[File:T--HUST-China--2018-tonglu-mles-lldp.png ‎|400px|thumb|center|Figure1. RBS-mleS-RBS-lldP]]
  
 
<h2>DNA Gel Electrophoretic</h2>
 
<h2>DNA Gel Electrophoretic</h2>
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To make sure that we get the target gene, we did the DNA gel electrophoretic to separate different gene. And here is the result.
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To make sure that we get the target gene, we did DNA gel electrophoretic for verification. And here is the result.
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[[File:T--HUST-China--2018-jiaotu-ldha and lldp.png|400px|thumb|center|Figure2:Verification of successful transformation of pSB1C3-RBS-ldhA]]
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[[File:T--HUST-China--2018-jiaotu-mles-lldp.png|400px|thumb|center|Figure2. Verification of successful transformation of pSB1C3-RBS-mleS-RBS-lldP]]
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Our target genes are 1047bp, and as the marker is DS5000, we could be sure that the bright bands in this picture are our target genes.
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Our target gene is 3323bp, and as the marker is DS5000, we could be sure that the bright bands in this picture are our target genes.
  
 
<h2>Electrogenesis</h2>
 
<h2>Electrogenesis</h2>
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By comparing the ability of producing electricity, we might find out whether dld could effectively help Shewanella to produce more electricity.
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By detecting the production of lactate after expressing, we might find out whether RBS-mleS-RBS-lldP could effectively help Rhodopseudomonas palustris transport produced lactate out of the cell.
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[[File:T--HUST-China--2018-expression of lactate.png ‎|400px|thumb|center|Figure3:shows that our modification is effective. Every gene circuits can help strains produce lactate, and mleS-lldP-ldhA is the most efficient one. Therefore, our construction of gene circuits achieve the goal to help strains produce lactate.]]
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[[File:T--HUST-China--2018-expression of lactate.png ‎|400px|thumb|center|Figure3. shows that our modification is effective. Every gene circuits can help strains produce lactate, and mleS-lldP-ldhA is the most efficient one. Therefore, our construction of gene circuits achieves the goal to help strains produce lactate.]]
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It could be demonstrated that targeted genes could be expressed in the engineered cells. More NADH has been produced by engineered bacteria, which helps to produce more electricty.  
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It could be demonstrated that the target genes could be expressed in the engineered cells. More lactate has been produced and transported by engineered bacteria.  
  
 
<!-- Uncomment this to enable Functional Parameter display  
 
<!-- Uncomment this to enable Functional Parameter display  

Latest revision as of 23:06, 17 October 2018


RBS-mleS-RBS-lldP

malate dehydrogenase & L-lactate permease

Usage and biology

It encodes malate dehydrogenase and L-lactate permease, which could produce and transport lactate. mleS could convert malic acid to L-lactate. lldP could transport lactate out of the cell. In this way, Rhodopseudomonas palustris could efficiently transport produced lactate out of the cell, which later will be used by Shewanella.

Characterization

This is one section for lactate production and transportation part.

Figure1. RBS-mleS-RBS-lldP

DNA Gel Electrophoretic

To make sure that we get the target gene, we did DNA gel electrophoretic for verification. And here is the result.

Figure2. Verification of successful transformation of pSB1C3-RBS-mleS-RBS-lldP

Our target gene is 3323bp, and as the marker is DS5000, we could be sure that the bright bands in this picture are our target genes.

Electrogenesis

By detecting the production of lactate after expressing, we might find out whether RBS-mleS-RBS-lldP could effectively help Rhodopseudomonas palustris transport produced lactate out of the cell.

Figure3. shows that our modification is effective. Every gene circuits can help strains produce lactate, and mleS-lldP-ldhA is the most efficient one. Therefore, our construction of gene circuits achieves the goal to help strains produce lactate.

It could be demonstrated that the target genes could be expressed in the engineered cells. More lactate has been produced and transported by engineered bacteria.