Difference between revisions of "Part:BBa K3552015"

 
Line 4: Line 4:
  
 
We improved the part BBa_K180000 to improve its productivity. We reversed the Lac operon parts and changed to a reverse promoter, terminator and RBS. A new ribozyme, RiboJ is added to the sequence. This part is in the part collection where we have 12 genes that code for the base generator of pilA.  
 
We improved the part BBa_K180000 to improve its productivity. We reversed the Lac operon parts and changed to a reverse promoter, terminator and RBS. A new ribozyme, RiboJ is added to the sequence. This part is in the part collection where we have 12 genes that code for the base generator of pilA.  
−
 
−
Our new ptac promoter consists of 7 components, the reverse lac terminator, lacI, RBS, lac promoter and normal ptac promoter, lacO optimized and RiboJ. The new promoter comparing to the BBa_K180000 found in 2009 has a much larger gradient of changing when comparing to the concentration of IPTG. In our experiment, we add different concentration of IPTG to constant promoter and received the value of florescence. We compared the value to the negative control and acquire the stable points to compare the data from the old ptac promoter.
 
−
 
−
<!-- Add more about the biology of this part here
 
−
===Usage and Biology===
 
  
 
<!-- -->
 
<!-- -->
Line 14: Line 9:
 
<partinfo>BBa_K3552015 SequenceAndFeatures</partinfo>
 
<partinfo>BBa_K3552015 SequenceAndFeatures</partinfo>
  
 +
==Characterization==
 +
Our new ptac promoter consists of 7 components, the reverse lac terminator, lacI, RBS, lac promoter and normal ptac promoter, lacO optimized and RiboJ. The new promoter comparing to the BBa_K180000 found in 2009 has a much larger gradient of changing when comparing to the concentration of IPTG. In our experiment, we add different concentration of IPTG to constant promoter and received the value of florescence. We compared the value to the negative control and acquire the stable points to compare the data from the old ptac promoter.
 +
 +
 +
The main carbon source is the glycerol and the bacteria were cultured in M9 medium in order to simulate the expression of pilA. In this way, the result might be able to become a reference relating to our pilA production. The graph shows that after adding RiboJ, SccJ, and SarJ, the tangent of the line increased remarkably, referring to an ascent of the E.coli sensitivity of IPTG in low concentration. RiboJ has the best function on increasing the productivity of GFP among the five ribozymes.
 +
We repeated the experiment in the culture medium with glucose as the source of carbon and gained similar results as the previous experiment. The data, in this manner, illustrate that the inducible promotor is robust in rising the yield in relatively low concentration. To our project, we have added RiboJ in front of our pilA genes to keep the gene in highly expression rate. In the following stage, we will apply these ribozymes on the generator transformation and better rise the production of pili generator. We hope this metamorphosis could generally increase the pili production.
  
 
<!-- Uncomment this to enable Functional Parameter display  
 
<!-- Uncomment this to enable Functional Parameter display  

Revision as of 04:08, 26 October 2020


Ptac-RiboJ

We improved the part BBa_K180000 to improve its productivity. We reversed the Lac operon parts and changed to a reverse promoter, terminator and RBS. A new ribozyme, RiboJ is added to the sequence. This part is in the part collection where we have 12 genes that code for the base generator of pilA.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]

Characterization

Our new ptac promoter consists of 7 components, the reverse lac terminator, lacI, RBS, lac promoter and normal ptac promoter, lacO optimized and RiboJ. The new promoter comparing to the BBa_K180000 found in 2009 has a much larger gradient of changing when comparing to the concentration of IPTG. In our experiment, we add different concentration of IPTG to constant promoter and received the value of florescence. We compared the value to the negative control and acquire the stable points to compare the data from the old ptac promoter.


The main carbon source is the glycerol and the bacteria were cultured in M9 medium in order to simulate the expression of pilA. In this way, the result might be able to become a reference relating to our pilA production. The graph shows that after adding RiboJ, SccJ, and SarJ, the tangent of the line increased remarkably, referring to an ascent of the E.coli sensitivity of IPTG in low concentration. RiboJ has the best function on increasing the productivity of GFP among the five ribozymes. We repeated the experiment in the culture medium with glucose as the source of carbon and gained similar results as the previous experiment. The data, in this manner, illustrate that the inducible promotor is robust in rising the yield in relatively low concentration. To our project, we have added RiboJ in front of our pilA genes to keep the gene in highly expression rate. In the following stage, we will apply these ribozymes on the generator transformation and better rise the production of pili generator. We hope this metamorphosis could generally increase the pili production.