Difference between revisions of "Team:UFlorida/Results"

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<h1>Results</h1>
 
<h1>Results</h1>
  
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<p>Liquid cultures of our bacteria were prepared in King’s Medium B supplemented with tryptophan. The culture was centrifuged after growth and the supernatant was filtered and used in HPLC. The figure shows tryptophol run in the same media as our sample. Our HPLC graphs do not have the peak at 162 m/z shown in the standard.
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<p>We transformed plasmids containing ARO10 in pETDUET-1 and ADH in pCOLADUET-1 in BL21 cells. We were unable to confirm the presence of ARO9 in a DUET vector either through gel electrophoresis or through Sanger Sequencing. ARO10 in pETDUET-1 and ADH in pCOLADUET-1 were validated using Sanger Sequencing by Genewiz®
 
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<p>From the HPLC results, the modified bacteria do not show production of tryptophol. One potential explanation is that there is post-translational modification of these gene products. In particular, ARO10 has been shown to be expressed without exhibiting 2-oxo acid decarboxylase activity, indicating the presence of post-translational modification that may be present in yeast cells but absent in the modified E. coli cells. </p>
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<p>We ran an HPLC Standard of tryptophol in King's Medium B. Tryptophol shows an elution peak at 162 m/z.  </p>
  
 
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<p>Most of our time over the summer was spent getting all three genes into one cell. In future experiments, PDC 1 could be used in place of ARO10 to produce the decarboxylase enzyme necessary for the degradation of tryptophan to tryptophol.</p>
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<p>We are planning to conduct HPLC using a filtered bacterial culture, and compare the graph generated to the graph of our tryptophol standard. We are planning on having HPLC results for or poster and presentation.</p>
 
   
 
   
  

Revision as of 23:41, 1 November 2017

Results

We transformed plasmids containing ARO10 in pETDUET-1 and ADH in pCOLADUET-1 in BL21 cells. We were unable to confirm the presence of ARO9 in a DUET vector either through gel electrophoresis or through Sanger Sequencing. ARO10 in pETDUET-1 and ADH in pCOLADUET-1 were validated using Sanger Sequencing by Genewiz®

We ran an HPLC Standard of tryptophol in King's Medium B. Tryptophol shows an elution peak at 162 m/z.

We are planning to conduct HPLC using a filtered bacterial culture, and compare the graph generated to the graph of our tryptophol standard. We are planning on having HPLC results for or poster and presentation.