Difference between revisions of "Team:Hamburg/Composite Part"

 
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<h1>Best Composite Part: <a href="http://parts.igemorg/Part:BBa_K2588021">BBa_K2588021</a></h1>
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                    <p>To create a Sustainable H.I.E.L.D. we needed the bacteria not to grow and divide, but to maintain a stable cell density. Our growth inhibition module <a href="http://parts.igemorg/Part:BBa_K2588021">BBa_K2588021</a> was confirmed by characterization. It inhibits <em>E. coli </em>cell division and growth to a third of its natural growth within only 6 hours after induction. <a href="http://parts.igemorg/Part:BBa_K2588021">BBa_K2588021</a> induces expression of growth inhibiting proteins cbtA, cspD and mraZ in absence of diaminopimelic acid (DAP). This allows to grow bacteria wearing this construct by addition of DAP but becomes active in natural environments without high amounts of DAP (Fig. 1).</p>
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                        <img src="https://static.igem.org/mediawiki/2018/0/04/T--Hamburg--GrowthInhibitionModule.svg" />
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                        <figcaption><b>Fig. 1: BBa_K2588021 Map.</b> DAP-repressed promoter dapAP induces expression of cell division inhibitors cbtA, cspD, and mraZ.</figcaption>
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                    <p>To characterize our growth inhibition module, we measured the optical density of the inhibited (+DAP), induced (-DAP), positive (DH5&alpha;) cultures at OD600 nm for 6 hours at 1-hour intervals. The part was assembled using the Golden Gate assembly method. The growth inhibition module is regulated by a dapAP promoter, which is inhibited by the presence of diaminopimelic acid. Measurement is shown in Fig. 2.&nbsp; The culture with (-DAP) had a reduced growth by factor 3.21 compared to the control. The culture with 40 &micro;M DAP added had a reduced growth by factor 2.30.&nbsp;&nbsp; </p>
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                        <img src="https://static.igem.org/mediawiki/2018/b/b3/T--Hamburg--GrowthInhibitionCharacterization.svg" />
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                        <figcaption><b>Fig. 2: Characterization of BBa_K2588021</b> Cultures containing the growth inhibition module were inhibit with DAP and OD<sub>600</sub> was measured for 7 h in 1 h intervals. Uninhibited cultures as well as DH5&alpha; were measured as positive controls.</figcaption>
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                    </figure>
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                    <p>Upon addition of DAP (+DAP), cultures containing the growth inhibition module showed a growth comparable to the positive control, while the induced growth inhibited culture (-DAP) exhibited weak growth. While we expected the induced growth inhibited culture to not proliferate and not growth, a increase in growth rate was observable. Therefore we confirm that the growth inhibition module indeed reduces cell proliferation.</p>
  
 
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<h1>Composite Parts</h1>
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<script type='text/javascript' src="https://2018.igem.org/wiki/index.php?title=Template:Hamburg/JS/main&action=raw&ctype=text/javascript"></script>
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A composite part is a functional unit of DNA consisting of two or more basic parts assembled together. <a href="http://parts.igem.org/wiki/index.php/Part:BBa_I13507">BBa_I13507</a> is an example of a composite part, consisting of an RBS, a protein coding region for a red fluorescent protein, and a terminator.
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<p>New composite BioBrick devices can be made by combining existing BioBrick Parts (like Inverters, Amplifiers, Smell Generators, Protein Balloon Generators, Senders, Receivers, Actuators, and so on).</p>
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<h3>Note</h3>
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<p>This page should list all the composite parts your team has made during your project. You must add all characterization information for your parts on the Registry. You should not put characterization information on this page. Remember judges will only look at the first part in the list for the Best Composite Part award, so put your best part first!</p>
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<h3>Best Composite Part Special Prize</h3>
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<p>To be eligible for this award, this part must adhere to <a href="http://parts.igem.org/DNA_Submission">Registry sample submission guidelines</a> and have been sent to the Registry of Standard Biological Parts. If you have a part you wish to nominate your team for this <a href="https://2018.igem.org/Judging/Awards">special prize</a>, make sure you add your part number to your <a href="https://2018.igem.org/Judging/Judging_Form">judging form</a> and delete the box at the top of this page.
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<b>Please note:</b> Judges will only look at the first part number you list, so please only enter ONE (1) part number in the judging form for this prize. </p>
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Latest revision as of 02:38, 18 October 2018

Best Composite Part: BBa_K2588021

To create a Sustainable H.I.E.L.D. we needed the bacteria not to grow and divide, but to maintain a stable cell density. Our growth inhibition module BBa_K2588021 was confirmed by characterization. It inhibits E. coli cell division and growth to a third of its natural growth within only 6 hours after induction. BBa_K2588021 induces expression of growth inhibiting proteins cbtA, cspD and mraZ in absence of diaminopimelic acid (DAP). This allows to grow bacteria wearing this construct by addition of DAP but becomes active in natural environments without high amounts of DAP (Fig. 1).

Fig. 1: BBa_K2588021 Map. DAP-repressed promoter dapAP induces expression of cell division inhibitors cbtA, cspD, and mraZ.

To characterize our growth inhibition module, we measured the optical density of the inhibited (+DAP), induced (-DAP), positive (DH5α) cultures at OD600 nm for 6 hours at 1-hour intervals. The part was assembled using the Golden Gate assembly method. The growth inhibition module is regulated by a dapAP promoter, which is inhibited by the presence of diaminopimelic acid. Measurement is shown in Fig. 2.  The culture with (-DAP) had a reduced growth by factor 3.21 compared to the control. The culture with 40 µM DAP added had a reduced growth by factor 2.30.  

Fig. 2: Characterization of BBa_K2588021 Cultures containing the growth inhibition module were inhibit with DAP and OD600 was measured for 7 h in 1 h intervals. Uninhibited cultures as well as DH5α were measured as positive controls.

Upon addition of DAP (+DAP), cultures containing the growth inhibition module showed a growth comparable to the positive control, while the induced growth inhibited culture (-DAP) exhibited weak growth. While we expected the induced growth inhibited culture to not proliferate and not growth, a increase in growth rate was observable. Therefore we confirm that the growth inhibition module indeed reduces cell proliferation.

Funding