Difference between revisions of "Part:BBa K216009:Experience"

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This part was designed to characterize the activity of the PyeaR promoter by Miller assay, as a backup to the GFP-based construct used in the promoter characterization kit.
 
This part was designed to characterize the activity of the PyeaR promoter by Miller assay, as a backup to the GFP-based construct used in the promoter characterization kit.
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<b><u>Results from Miller's Assay</u></b>
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<br>
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[[Image:PyeaR-miller.png]]
  
 
===User Reviews===
 
===User Reviews===
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'''Initial Tests: Edinburgh iGEM 2009:''' To test this part, we grew ''E. coli'' JM109 overnight at 37 C with shaking in LB with 80 mg/l ampicillin and varying concentrations of sodium nitrate and sodium nitrite. The following day, we performed Miller assays (LacZ activity assays using o-nitrophenyl galactoside, ONPG, as a chromogenic substrate). The results showed strong induction by both nitrate and nitrite, as expected for this promoter (see biology information for part BBa_K216005).
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'''Further Tests: BCCS-Bristol iGEM 2010:'''
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*Cultures of ''E.coli'' strain XL1-Blue transformed with BBa_K216009 were grown at 0mM, 15mM and 30mM [KNO<sub>3</sub>]. Two parallel sets of cultures were set up, one in the presence of IPTG (set 1), one in the absence of IPTG (set 2). All cultures were grown in both Ampicillin and Tetracyclin together. We then performed a β-galactosidase assay on the cultures, with the following results
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*Table
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[[Image:BCCS_attempt3betagaltable.jpg|frameless|center|upright=4|table of betagal results]]
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*Graph
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[[Image:BCCS attempt3betagalgraph.jpg|frameless|center|upright=4|graph of betagal results]]
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<!-- Template for a user review
 
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Latest revision as of 18:53, 27 October 2010

This experience page is provided so that any user may enter their experience using this part.
Please enter how you used this part and how it worked out.

Applications of BBa_K216009

This part was designed to characterize the activity of the PyeaR promoter by Miller assay, as a backup to the GFP-based construct used in the promoter characterization kit.

Results from Miller's Assay
PyeaR-miller.png

User Reviews

Initial Tests: Edinburgh iGEM 2009: To test this part, we grew E. coli JM109 overnight at 37 C with shaking in LB with 80 mg/l ampicillin and varying concentrations of sodium nitrate and sodium nitrite. The following day, we performed Miller assays (LacZ activity assays using o-nitrophenyl galactoside, ONPG, as a chromogenic substrate). The results showed strong induction by both nitrate and nitrite, as expected for this promoter (see biology information for part BBa_K216005).

Further Tests: BCCS-Bristol iGEM 2010:

  • Cultures of E.coli strain XL1-Blue transformed with BBa_K216009 were grown at 0mM, 15mM and 30mM [KNO3]. Two parallel sets of cultures were set up, one in the presence of IPTG (set 1), one in the absence of IPTG (set 2). All cultures were grown in both Ampicillin and Tetracyclin together. We then performed a β-galactosidase assay on the cultures, with the following results


  • Table
table of betagal results


  • Graph
graph of betagal results

UNIQ3018ce4ae41d3857-partinfo-00000000-QINU UNIQ3018ce4ae41d3857-partinfo-00000001-QINU