Difference between revisions of "Part:BBa K540000:Experience"
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===Applications of BBa_K540000===
 
===Applications of BBa_K540000===
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<html>
 
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<div style="text-align:center; float:right; margin-left:30px; margin-top:-20px;">
 
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<img src="http://i.imgur.com/g7iTW.png" style="width:340px; height:255px; margin-top: 70px; margin-bottom: 5px; position: relative;"/>
 
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</div>
 
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<p>
 
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A preliminary search on the best condition for use was carried out at first.
 
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</p><br>
 
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<p>
 
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<FONT COLOR="green"><b>Antibiotic response</b></FONT><br>
 
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Two 24 wells plates have been seeded with the same strains, in LB/2 medium. Ampicilline for S18 and Cm have been added only in the second plate.
 
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The results have examined after 24 hours of incubation 30°C.
 
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</p><br>
 
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<p><i>Qualitative approach</i><br>
 
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As can be seen on the right, adding ampicilline boosts the adherence of the strain, and makes it more cobalt responsive.
 
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</p>
 
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<div style="text-align:center; float:left; margin-left:30px; margin-top:-20px;">
 
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<img src="http://i.imgur.com/wnOY5.png" style="width:525px; height:350px; margin-top: 100px; margin-bottom: 5px; position: relative;"/>
 
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</div>
 
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<p>
 
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<br/><br/>
 
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<i>Quantitative approach</i><br>
 
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Furthermore, the adherence level was determined quantitatively using OD measurements. Resulting graphs are shown on the left.</p><br>
 
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<p>
 
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P1 indicates the first plate, without antibiotics, while P2 indicates the second plate, with antibiotics.</p><br>
 
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<p>
 
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<b>Conclusion</b>: it can clearly be seen that while the growth of the bacteria are similar, as they have similar total OD, adding the antibiotics makes them a lot more adherent, and enable the cobalt-response.</p><br>
 
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<p>
 
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This can be explained by the fact that without antibiotics, the bacteria will tend to lose the part, and thus become unresponsive to cobalt.
 
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</p><br>
 
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<div style="text-align:center; float:right; margin-left:30px; margin-top:-20px;">
 
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<img src="http://i.imgur.com/zIwBU.png" style="width:402px; height:350px; margin-top: 60px; margin-bottom: 5px; position: relative;"/>
 
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</div>
 
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<br><br><br><br>
 
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<p>
 
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<FONT COLOR="green"><b>Medium influence</b></FONT><br>
 
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During adherence tests, bacteria can be grown in LB/2 medium (50% Lb, 50% sterile water) or in M63G medium (100mL M63, 1ml LB, 1 ml Glucose 20%).
 
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</p><br>
 
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<p>
 
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In order to choose the better medium, two strains have been seeded in the 2 different media, with a concentration of 10^7 cells/mL. The first is an adherent strain PHL818, the second is a non-adherent strain NM522.
 
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Plates have been incubated at 30°C during 48 hours.
 
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</p><br>
 
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<p>
 
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Results which are show on the right are a compilation of repetitions of this experiment.
 
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</p><br>
 
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<p>
 
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<b>Conclusion</b>: a greater biofilm formation is observed using the M63G medium.
 
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<div style="text-align:center; float:left; margin-right:20px; margin-top:-20px;">
 
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<img src="http://i.imgur.com/p0pEe.png" style="width:308px; height:447px; margin-top: 110px; margin-bottom: 5px; position: relative;"/>
 
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</div>
 
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<br><br><br><br><br>
 
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<p>
 
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<FONT COLOR="green"><b>EDTA cleaning influence</b></FONT><br>
 
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Plates were washed with 4mM EDTA, and rinsed with water before growing bacteria. This aims to reduce the influence of any cobalt or nickel pre-existing in the plates, that might change the actual cobalt concentration in the plates.
 
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</p><br>
 
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<p>
 
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Results are on the left. As can be seen, treating with EDTA beforehand increases the adherence of the strain.
 
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</p><br>
 
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<p>
 
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<b>Conclusion</b>: When using this part, it is preferable to wash the plates beforehand with EDTA (or to use metal-free plastic)
 
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</p>
 
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<br/><br/><br/><br/><br><br><br><br>
 
  
 
===User Reviews===
 
===User Reviews===
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<partinfo>BBa_K540000 AddReview number</partinfo>
 
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<I>Username</I>
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<I>User</I>
 
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Enter the review inofrmation here.
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<partinfo>BBa_I0500 AddReview 5</partinfo>
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<I>UCL iGEM 2012</I>
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UCL iGEM 2012 have removed the original cobalt inducible promoter to create a stand alone csgBAEFG operon with rbs [https://parts.igem.org/wiki/index.php?title=Part:BBa_K729003 BBa_K729003]. We have also designed two additional variations on the part, with a [https://parts.igem.org/wiki/index.php?title=Part:BBa_K729017 T7] and [https://parts.igem.org/wiki/index.php?title=Part:BBa_K729018 constitutive promoter].
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Latest revision as of 19:30, 28 September 2012

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_K540000

User Reviews

UNIQe4ef173237708238-partinfo-00000000-QINU UNIQe4ef173237708238-partinfo-00000001-QINU

•••••

UCL iGEM 2012

UCL iGEM 2012 have removed the original cobalt inducible promoter to create a stand alone csgBAEFG operon with rbs BBa_K729003. We have also designed two additional variations on the part, with a T7 and constitutive promoter.