Difference between revisions of "Part:BBa K2963033"

 
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<partinfo>BBa_K2963033 short</partinfo>
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<partinfo>BBa_K2963032 short</partinfo>
  
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This part contains the racE gene for glutamate racemase from Bacillus. This enzyme mainly catelyse the reation of L-Glumate to D-Glumate.In our project, we express this enzyme by using the "Ptac-lacO-lacO-rbs-racE-T7t" part in order to reduce the leaked expression of racE.
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This part contains the racE racemase gene, which was constructed using the tac (BBa_K864400) promoter, rbs (BBa_K2963006), racE gene (BBa_K2963004) and the T7 terminator (BBa_K2598024). This part was compared with the other part (BBa_K2963033) which contained another lac operator follwowing the tac promoter（BBa_K864400）.Simultaneously, the rest basic parts are the same. The racemase activity data and real-time PCR data were used to characterize the composite part. And we compared the different expression levels of racE gene of the two composite parts(BBa_K2963033 and BBa_K2963032）.
  
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===Usage and Biology===
 
===Usage and Biology===
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The racE gene is a type of racemase gene from Bacillus subtilis while in Bacillus licheniformis is named glr. According to previous reports, glr has two characters against L-glutamic acid, one is weak affinity and the other is strong catalytic properties.
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===Characterization===
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[[image:Figure1-gaohaixin.png|400px]]
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Figure 1. The expression level of the racemase gene racE at 8h,16h,24h decreased by 29.99%, 58.86% and 62.34% respectively. The results show that the expression intensity of racE was effectively reduced at each transcriptional stage by the tandem of two lacOs compared to one lacO.
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[[image:Figure2-gaohaixin.png|400px]]
  
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Figure 2. We tested the enzyme activity of the racemase in cells. The result was the same as the above and the enzyme activity was different.
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<span class='h3bb'>Sequence and Features</span>
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<partinfo>BBa_K2963033 SequenceAndFeatures</partinfo>
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===Functional Parameters===
 
===Functional Parameters===
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<partinfo>BBa_K2963033 parameters</partinfo>
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<partinfo>BBa_K2963032 parameters</partinfo>
 
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Revision as of 15:20, 13 October 2019

racE- encoding racemase

This part contains the racE racemase gene, which was constructed using the tac (BBa_K864400) promoter, rbs (BBa_K2963006), racE gene (BBa_K2963004) and the T7 terminator (BBa_K2598024). This part was compared with the other part (BBa_K2963033) which contained another lac operator follwowing the tac promoter（BBa_K864400）.Simultaneously, the rest basic parts are the same. The racemase activity data and real-time PCR data were used to characterize the composite part. And we compared the different expression levels of racE gene of the two composite parts(BBa_K2963033 and BBa_K2963032）.

Usage and Biology

The racE gene is a type of racemase gene from Bacillus subtilis while in Bacillus licheniformis is named glr. According to previous reports, glr has two characters against L-glutamic acid, one is weak affinity and the other is strong catalytic properties.

Characterization

Figure1-gaohaixin.png

Figure 1. The expression level of the racemase gene racE at 8h,16h,24h decreased by 29.99%, 58.86% and 62.34% respectively. The results show that the expression intensity of racE was effectively reduced at each transcriptional stage by the tandem of two lacOs compared to one lacO.

Figure2-gaohaixin.png

Figure 2. We tested the enzyme activity of the racemase in cells. The result was the same as the above and the enzyme activity was different.