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content.titleLong = "A list of generated plasmids"

content.summary= "A list of each and every plasmid we designed combined with short summaries of their function";

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content.text="<a href='https://static.igem.org/mediawiki/2013/e/ef/Bonn_AraC-pBAD%28D%29-RBS32-mazF-TT_BBa_K1075040.jpg'><img src='https://static.igem.org/mediawiki/2013/e/ef/Bonn_AraC-pBAD%28D%29-RBS32-mazF-TT_BBa_K1075040.jpg' width='840' height='200'></a>MazF is part of the biological toxin-antitoxin system MazEF. This system is derived from the bacterium Escherichia coli, where it serves as a stress inducable killswitch. MazF, a RNA-degrading enzyme, ist he toxin and is normally inhibited by the presence of MazE. Absence of MazE leads to cell death.<a href=http://parts.igem.org/Part:BBa_K1075040>1</a><a href='https://static.igem.org/mediawiki/2013/3/34/Bonn_pLac1-RBS32-mazF-TT_BBa_K1075039.jpg'><img src='https://static.igem.org/mediawiki/2013/3/34/Bonn_pLac1-RBS32-mazF-TT_BBa_K1075039.jpg' width='840' height='200'></a>We used this part to gain an IPTG and Lactose inducible construct of MazF. This part was used to test the toxicity of MazF as part of our proof of concept. <a href=http://parts.igem.org/Part:BBa_K1075039>2</a><a href='https://static.igem.org/mediawiki/2013/8/87/Bonn_pLac2-RBS32-mazE-%28ec%29ssrA%28DAS%2B4%29-TT_BBa_K1075043.jpg'><img src='https://static.igem.org/mediawiki/2013/8/87/Bonn_pLac2-RBS32-mazE-%28ec%29ssrA%28DAS%2B4%29-TT_BBa_K1075043.jpg' width='840' height='200'></a>This part can be used to gain a Lactose and IPTG inducable construct of mazE-(ec)ssrA(DAS+4). We used this construct as part of our proof of concept. <a href=http://parts.igem.org/Part:BBa_K1075043>2</a>";

+

content.text="<a href='https://static.igem.org/mediawiki/2013/e/ef/Bonn_AraC-pBAD%28D%29-RBS32-mazF-TT_BBa_K1075040.jpg'><img src='https://static.igem.org/mediawiki/2013/e/ef/Bonn_AraC-pBAD%28D%29-RBS32-mazF-TT_BBa_K1075040.jpg' width='840' height='200'></a>MazF is part of the biological toxin-antitoxin system MazEF. This system is derived from the bacterium Escherichia coli, where it serves as a stress inducable killswitch. MazF, a RNA-degrading enzyme, ist he toxin and is normally inhibited by the presence of MazE. Absence of MazE leads to cell death.<a href=http://parts.igem.org/Part:BBa_K1075040>1</a><a href='https://static.igem.org/mediawiki/2013/3/34/Bonn_pLac1-RBS32-mazF-TT_BBa_K1075039.jpg'><img src='https://static.igem.org/mediawiki/2013/3/34/Bonn_pLac1-RBS32-mazF-TT_BBa_K1075039.jpg' width='840' height='200'></a>We used this part to gain an IPTG and Lactose inducible construct of MazF. This part was used to test the toxicity of MazF as part of our proof of concept. <a href=http://parts.igem.org/Part:BBa_K1075039>2</a><a href='https://static.igem.org/mediawiki/2013/8/87/Bonn_pLac2-RBS32-mazE-%28ec%29ssrA%28DAS%2B4%29-TT_BBa_K1075043.jpg'><img src='https://static.igem.org/mediawiki/2013/8/87/Bonn_pLac2-RBS32-mazE-%28ec%29ssrA%28DAS%2B4%29-TT_BBa_K1075043.jpg' width='840' height='200'></a>This part can be used to gain a Lactose and IPTG inducable construct of mazE-(ec)ssrA(DAS+4). We used this construct as part of our proof of concept. <a href=http://parts.igem.org/Part:BBa_K1075043>3</a><a href='https://static.igem.org/mediawiki/2013/c/cf/Bonn_AraC-pBAD-RBS-mazF-TT-pLac2-RBS-mazE-%28ec%29ssrA%28DAS%2B4%29-TT_BBa_K1075048.jpg'><img src='https://static.igem.org/mediawiki/2013/c/cf/Bonn_AraC-pBAD-RBS-mazF-TT-pLac2-RBS-mazE-%28ec%29ssrA%28DAS%2B4%29-TT_BBa_K1075048.jpg' width='840' height='200'></a>This construct can be used to gain IPTG and Lactose inducible expression of mazE(ec)ssrA(DAS+4) and arabinose inducible expression of MazF. Thus, both parts of the toxin-antitoxin system mazEF are inducible and this construct can be used to further examine this system. We used this construct as part of our proof of concept.<a href=http://parts.igem.org/Part:BBa_K1075048>4</a><a href='https://static.igem.org/mediawiki/2013/7/7f/Bonn_AraC-pBAD-RBS-sspB-TT-pLac2-RBS-mCherry-TT_BBa_K1075049.jpg'><img src='https://static.igem.org/mediawiki/2013/7/7f/Bonn_AraC-pBAD-RBS-sspB-TT-pLac2-RBS-mCherry-TT_BBa_K1075049.jpg' width='840' height='200'></a>The sspB protein is an adaptor responsible for delivering ssrA-tagged substrates to the ClpXP protease in order to enhance their degradation. mCherry is a red fluorescent protein with the excitation maximum at 587 nm and the Emission maximum at 610 nm.<a href=http://parts.igem.org/Part:BBa_K1075049>5</a><a href='https://static.igem.org/mediawiki/2013/4/42/Bonn_pDawn_%28Prom_%28const.%29-RBS34-YF1-FixJ-FixK-LambdaC-pC%29_BBa_K1075044.jpg'><img src='https://static.igem.org/mediawiki/2013/4/42/Bonn_pDawn_%28Prom_%28const.%29-RBS34-YF1-FixJ-FixK-LambdaC-pC%29_BBa_K1075044.jpg' width='840' height='200'></a>The plasmid pDawn was designed by Ohlendorf et al. in 2012 together with its counter plasmid pDusk. Both plasmids are single plasmid systems, which allow the activation (pDawn) or repression (pDusk) of gene expression by blue light. They are easy to implement in the laboratory and lead to up to 460-fold activity change upon ilumination. <a href=http://parts.igem.org/Part:BBa_K1075044>6</a><a href='https://static.igem.org/mediawiki/2013/5/50/Bonn_pDawn_%28Prom_%28const.%29-RBS34-YF1-FixJ-FixK-LambdaC-pC%29-RBS32-ccdB-TT_BBa_K1075047.jpg'><img src='https://static.igem.org/mediawiki/2013/5/50/Bonn_pDawn_%28Prom_%28const.%29-RBS34-YF1-FixJ-FixK-LambdaC-pC%29-RBS32-ccdB-TT_BBa_K1075047.jpg' width='840' height='200'></a>pDawn-RBS32-ccdB-TT can be used to photocontrol the expression of the toxin ccdB. In this system pDawn (BBa_K1075044) can be regulated by blue light, while ccdB (BBa_K1075029) acts as a killswitch. The ccd operon of the F plasmid encodes CcdB, a toxin targeting the essential gyrase of Escherichia coli, and CcdA, the unstable antidote that interacts with CcdB to neutralize its toxicity.<a href=http://parts.igem.org/Part:BBa_K1075047>7</a><a href='https://static.igem.org/mediawiki/2013/0/03/Bonn_AraC-pBAD%28D%29-RBS32-SspB%28Core%28-LOV-ipaA-TT_BBa_K1075019.jpg'><img src='https://static.igem.org/mediawiki/2013/0/03/Bonn_AraC-pBAD%28D%29-RBS32-SspB%28Core%28-LOV-ipaA-TT_BBa_K1075019.jpg' width='840' height='200'></a>The part can be used to regulate the dimerization of the two parts of the split sspB. The binding of ipaA to Vincolin is regulated via the light-sensitive LOV domain. Therefore the two sspB parts only come together when the LOV domain was activated with blue light. <a href=http://parts.igem.org/Part:BBa_K1075019>8</a><a href='https://static.igem.org/mediawiki/2013/d/d0/Bonn_J23105-RBS32-SspB%28Core%29-LOV-ipaA-TT_BBa_K1075018.jpg'><img src='https://static.igem.org/mediawiki/2013/d/d0/Bonn_J23105-RBS32-SspB%28Core%29-LOV-ipaA-TT_BBa_K1075018.jpg' width='840' height='200'></a>The part can be used to regulate the dimerization of the two parts of the split sspB. The binding of ipaA to Vincolin is regulated via the light-sensitive LOV domain. Therefore the two sspB parts only come together when the LOV domain was activated with blue light. <a href=http://parts.igem.org/Part:BBa_K1075018>9</a><a href='https://static.igem.org/mediawiki/2013/5/5a/Bonn_AraC-pBAD%28D%29-RBS32-VinD1-sspBXB-TT_BBa_K1075016.jpg'><img src='https://static.igem.org/mediawiki/2013/5/5a/Bonn_AraC-pBAD%28D%29-RBS32-VinD1-sspBXB-TT_BBa_K1075016.jpg' width='840' height='200'></a>The part can be used to regulate the dimerization of the two parts of the split sspB. As Vincolin bins to ipaA, the sspB dimer is stabilized. We used this part to achieve a light-induced dimerization of the sspB parts. <a href=http://parts.igem.org/Part:BBa_K1075016>10</a><a href='https://static.igem.org/mediawiki/2013/9/9c/Bonn_J23118-RBS32-VinD1-SspBXB-TT_BBa_K1075015.jpg'><img src='https://static.igem.org/mediawiki/2013/9/9c/Bonn_J23118-RBS32-VinD1-SspBXB-TT_BBa_K1075015.jpg' width='840' height='200'></a>The part can be used to regulate the dimerization of the two parts of the split sspB. As Vincolin bins to ipaA, the sspB dimer is stabilized. We used this part to achieve a light-induced dimerization of the sspB parts. <a href=http://parts.igem.org/Part:BBa_K1075015>11</a><a href='https://static.igem.org/mediawiki/2013/7/78/Bonn_pLac_pBAD2-RBS32-SspB-TT_BBa_K1075006.jpg'><img src='https://static.igem.org/mediawiki/2013/7/78/Bonn_pLac_pBAD2-RBS32-SspB-TT_BBa_K1075006.jpg' width='840' height='200'></a>The sspB protein is an adaptor responsible for delivering ssrA-tagged substrates to the ClpXP protease in order to enhance their degradation. Prefixed is RBS, the binding site for a ribosome and the pLac promoter. It also includes a double terminator 'TT', which interrupts the translation. <a href=http://parts.igem.org/Part:BBa_K1075006>12</a><a href='https://static.igem.org/mediawiki/2013/8/8f/Bonn_E._coli_ssrA%28DAS%2B4%29-TT_BBa_K1075021.jpg'><img src='https://static.igem.org/mediawiki/2013/8/8f/Bonn_E._coli_ssrA%28DAS%2B4%29-TT_BBa_K1075021.jpg' width='840' height='200'></a>The (ec)ssrA(DAS+4) tag can be fused to proteins so that they will be delivered to the ClpXP protease and degraded.<a href=http://parts.igem.org/Part:BBa_K1075021>13</a><a href='https://static.igem.org/mediawiki/2013/6/63/Bonn_AraC-pBad-RBS34-mCherry-ecssrA%28DAS%2B4%29-TT_BBa_K1075025.jpg'><img src='https://static.igem.org/mediawiki/2013/6/63/Bonn_AraC-pBad-RBS34-mCherry-ecssrA%28DAS%2B4%29-TT_BBa_K1075025.jpg' width='840' height='200'></a>As we want to control protein degradation by controlling the function of ecSspB, we tagged the red fluorescent protein mCherry with ecssrA(DAS+4) to measure the degradation rate. The part was designed for proof of principle. Application as a bacterial fotographic film might be possible as well. <a href=http://parts.igem.org/Part:BBa_K1075025>14</a><a href='https://static.igem.org/mediawiki/2013/6/63/Bonn_AraC-pBad-RBS34-mCherry-ecssrA%28DAS%2B4%29-TT_BBa_K1075025.jpg'><img src='https://static.igem.org/mediawiki/2013/6/63/Bonn_AraC-pBad-RBS34-mCherry-ecssrA%28DAS%2B4%29-TT_BBa_K1075025.jpg' width='840' height='200'></a>As we want to control protein degradation by controlling the function of ecSspB, we tagged the red fluorescent protein mCherry with ecssrA(DAS+4) to measure the degradation rate. The part was designed for proof of principle. Application as a bacterial fotographic film might be possible as well. <a href=http://parts.igem.org/Part:BBa_K1075025>15</a><a href='https://static.igem.org/mediawiki/2013/c/cd/Bonn_pJD427%28pC-RBS32-FKBP12-EcsspB%28XB%29-Term-pB-RBS32-EcsspB%28Core%29-FRB-Term-Term%29_BBa_K1075010.jpg'><img src='https://static.igem.org/mediawiki/2013/c/cd/Bonn_pJD427%28pC-RBS32-FKBP12-EcsspB%28XB%29-Term-pB-RBS32-EcsspB%28Core%29-FRB-Term-Term%29_BBa_K1075010.jpg' width='840' height='200'></a>The SspB protein is an adaptor responsible for delivering ssrA-tagged substrates to the ClpXP protease in order to enhance their degradation. To control degradation it is reasonable to control the function of SspB. That is made by splitting it into two parts, each of which cannot induce degradation on its own. To bring both SspB parts together again for inducible degradation, they were combined with a chemical inducible heterodimerisation system: FRB and FKBP12. These two parts interact in the presence of rapamycin <a href=http://parts.igem.org/Part:BBa_K1075010>16</a><a href='https://static.igem.org/mediawiki/2013/d/d6/Bonn_%28pJD427%29-pLac-RBS34-mCherry-ecssrA%28DAS%2B4%29-TT_BBa_K1075027.jpg'><img src='https://static.igem.org/mediawiki/2013/d/d6/Bonn_%28pJD427%29-pLac-RBS34-mCherry-ecssrA%28DAS%2B4%29-TT_BBa_K1075027.jpg' width='840' height='200'></a>As we want to control protein degradation by controlling the function of ecSspB, we tag the red fluorescent protein mCherry with ecssrA(DAS+4) to measure the degradation rate. We control ecSspB by splitting it into two parts each of which cannot induce degradation on its own but regains function by addition of rapamycin. As the co-transformation of the two plasmids didn’t work, we combined them to one.<a href=http://parts.igem.org/Part:BBa_K1075027>17</a><a href='https://static.igem.org/mediawiki/2013/b/bc/Bonn_pJD427-pLac-RBS34-mCherry-TT_BBa_K1075028.jpg'><img src='https://static.igem.org/mediawiki/2013/b/bc/Bonn_pJD427-pLac-RBS34-mCherry-TT_BBa_K1075028.jpg' width='840' height='200'></a>This plasmid is a negative control to the pJD427-pLac2-RBS34-mCherry-ecssrA(DAS+4)-TT plasmid. <a href=http://parts.igem.org/Part:BBa_K1075028>18</a><a href='https://static.igem.org/mediawiki/2013/c/cd/Bonn_pJD427%28pC-RBS32-FKBP12-EcsspB%28XB%29-Term-pB-RBS32-EcsspB%28Core%29-FRB-Term-Term%29_BBa_K1075010.jpg'><img src='https://static.igem.org/mediawiki/2013/c/cd/Bonn_pJD427%28pC-RBS32-FKBP12-EcsspB%28XB%29-Term-pB-RBS32-EcsspB%28Core%29-FRB-Term-Term%29_BBa_K1075010.jpg' width='840' height='200'></a>The Plasmid pJD427 contains the fusion proteins SspB[CORE]-FRB and FKBP12-SspB[XB]: SspB[CORE]-FRB with the weak constitutive promoter proB, FKBP12-SspB[XB] with the strong constitutive promoter proC and a medium-copy p15a origin of replication. <a href=http://parts.igem.org/Part:BBa_K1075010>19</a><a href='https://static.igem.org/mediawiki/2013/e/e1/Bonn_pLac-RBS32-ccdA-ssrA-TT_BBa_K1075035.jpg'><img src='https://static.igem.org/mediawiki/2013/e/e1/Bonn_pLac-RBS32-ccdA-ssrA-TT_BBa_K1075035.jpg' width='840' height='200'></a>CcdA inhibits the toxin ccdB by binding to it and thus represses cell death. The part contains ccdA with a ssrA tag under the control of the Lac operon. <a href=http://parts.igem.org/Part:BBa_K1075035>20</a><a href='https://static.igem.org/mediawiki/2013/8/81/Bonn_AraC-pBAD%28D%29-RBS32-ccdB-TT_BBa_K1075031.jpg'><img src='https://static.igem.org/mediawiki/2013/8/81/Bonn_AraC-pBAD%28D%29-RBS32-ccdB-TT_BBa_K1075031.jpg' width='840' height='200'></a>The part contains a chemically induced kill-switch. When Arabinose is added to the bacteria, the toxin ccdB is expressed and cell death is initiated. <a href=http://parts.igem.org/Part:BBa_K1075031>20</a>";

content.type="Project";

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@@ Line 643: / Line 643: @@
 content.titleLong = "A list of generated plasmids"
 content.summary= "A list of each and every plasmid we designed combined with short summaries of their function";
-content.text="<a href='https://static.igem.org/mediawiki/2013/e/ef/Bonn_AraC-pBAD%28D%29-RBS32-mazF-TT_BBa_K1075040.jpg'><img src='https://static.igem.org/mediawiki/2013/e/ef/Bonn_AraC-pBAD%28D%29-RBS32-mazF-TT_BBa_K1075040.jpg' width='840' height='200'></a>MazF is part of the biological toxin-antitoxin system MazEF. This system is derived from the bacterium Escherichia coli, where it serves as a stress inducable killswitch. MazF, a RNA-degrading enzyme, ist he toxin and is normally inhibited by the presence of MazE. Absence of MazE leads to cell death.<sup><a href=http://parts.igem.org/Part:BBa_K1075040>1</a></sup><a href='https://static.igem.org/mediawiki/2013/3/34/Bonn_pLac1-RBS32-mazF-TT_BBa_K1075039.jpg'><img src='https://static.igem.org/mediawiki/2013/3/34/Bonn_pLac1-RBS32-mazF-TT_BBa_K1075039.jpg' width='840' height='200'></a>We used this part to gain an IPTG and Lactose inducible construct of MazF. This part was used to test the toxicity of MazF as part of our proof of concept. <sup><a href=http://parts.igem.org/Part:BBa_K1075039>2</a></sup><a href='https://static.igem.org/mediawiki/2013/8/87/Bonn_pLac2-RBS32-mazE-%28ec%29ssrA%28DAS%2B4%29-TT_BBa_K1075043.jpg'><img src='https://static.igem.org/mediawiki/2013/8/87/Bonn_pLac2-RBS32-mazE-%28ec%29ssrA%28DAS%2B4%29-TT_BBa_K1075043.jpg' width='840' height='200'></a>This part can be used to gain a Lactose and IPTG inducable construct of mazE-(ec)ssrA(DAS+4). We used this construct as part of our proof of concept. <sup><a href=http://parts.igem.org/Part:BBa_K1075043>2</a></sup>";
+content.text="<a href='https://static.igem.org/mediawiki/2013/e/ef/Bonn_AraC-pBAD%28D%29-RBS32-mazF-TT_BBa_K1075040.jpg'><img src='https://static.igem.org/mediawiki/2013/e/ef/Bonn_AraC-pBAD%28D%29-RBS32-mazF-TT_BBa_K1075040.jpg' width='840' height='200'></a>MazF is part of the biological toxin-antitoxin system MazEF. This system is derived from the bacterium Escherichia coli, where it serves as a stress inducable killswitch. MazF, a RNA-degrading enzyme, ist he toxin and is normally inhibited by the presence of MazE. Absence of MazE leads to cell death.<sup><a href=http://parts.igem.org/Part:BBa_K1075040>1</a></sup><a href='https://static.igem.org/mediawiki/2013/3/34/Bonn_pLac1-RBS32-mazF-TT_BBa_K1075039.jpg'><img src='https://static.igem.org/mediawiki/2013/3/34/Bonn_pLac1-RBS32-mazF-TT_BBa_K1075039.jpg' width='840' height='200'></a>We used this part to gain an IPTG and Lactose inducible construct of MazF. This part was used to test the toxicity of MazF as part of our proof of concept. <sup><a href=http://parts.igem.org/Part:BBa_K1075039>2</a></sup><a href='https://static.igem.org/mediawiki/2013/8/87/Bonn_pLac2-RBS32-mazE-%28ec%29ssrA%28DAS%2B4%29-TT_BBa_K1075043.jpg'><img src='https://static.igem.org/mediawiki/2013/8/87/Bonn_pLac2-RBS32-mazE-%28ec%29ssrA%28DAS%2B4%29-TT_BBa_K1075043.jpg' width='840' height='200'></a>This part can be used to gain a Lactose and IPTG inducable construct of mazE-(ec)ssrA(DAS+4). We used this construct as part of our proof of concept. <sup><a href=http://parts.igem.org/Part:BBa_K1075043>3</a></sup><a href='https://static.igem.org/mediawiki/2013/c/cf/Bonn_AraC-pBAD-RBS-mazF-TT-pLac2-RBS-mazE-%28ec%29ssrA%28DAS%2B4%29-TT_BBa_K1075048.jpg'><img src='https://static.igem.org/mediawiki/2013/c/cf/Bonn_AraC-pBAD-RBS-mazF-TT-pLac2-RBS-mazE-%28ec%29ssrA%28DAS%2B4%29-TT_BBa_K1075048.jpg' width='840' height='200'></a>This construct can be used to gain IPTG and Lactose inducible expression of mazE(ec)ssrA(DAS+4) and arabinose inducible expression of MazF. Thus, both parts of the toxin-antitoxin system mazEF are inducible and this construct can be used to further examine this system. We used this construct as part of our proof of concept.<sup><a href=http://parts.igem.org/Part:BBa_K1075048>4</a></sup><a href='https://static.igem.org/mediawiki/2013/7/7f/Bonn_AraC-pBAD-RBS-sspB-TT-pLac2-RBS-mCherry-TT_BBa_K1075049.jpg'><img src='https://static.igem.org/mediawiki/2013/7/7f/Bonn_AraC-pBAD-RBS-sspB-TT-pLac2-RBS-mCherry-TT_BBa_K1075049.jpg' width='840' height='200'></a>The sspB protein is an adaptor responsible for delivering ssrA-tagged substrates to the ClpXP protease in order to enhance their degradation. mCherry is a red fluorescent protein with the excitation maximum at 587 nm and the Emission maximum at 610 nm.<sup><a href=http://parts.igem.org/Part:BBa_K1075049>5</a></sup><a href='https://static.igem.org/mediawiki/2013/4/42/Bonn_pDawn_%28Prom_%28const.%29-RBS34-YF1-FixJ-FixK-LambdaC-pC%29_BBa_K1075044.jpg'><img src='https://static.igem.org/mediawiki/2013/4/42/Bonn_pDawn_%28Prom_%28const.%29-RBS34-YF1-FixJ-FixK-LambdaC-pC%29_BBa_K1075044.jpg' width='840' height='200'></a>The plasmid pDawn was designed by Ohlendorf et al. in 2012 together with its counter plasmid pDusk. Both plasmids are single plasmid systems, which allow the activation (pDawn) or repression (pDusk) of gene expression by blue light. They are easy to implement in the laboratory and lead to up to 460-fold activity change upon ilumination. <sup><a href=http://parts.igem.org/Part:BBa_K1075044>6</a></sup><a href='https://static.igem.org/mediawiki/2013/5/50/Bonn_pDawn_%28Prom_%28const.%29-RBS34-YF1-FixJ-FixK-LambdaC-pC%29-RBS32-ccdB-TT_BBa_K1075047.jpg'><img src='https://static.igem.org/mediawiki/2013/5/50/Bonn_pDawn_%28Prom_%28const.%29-RBS34-YF1-FixJ-FixK-LambdaC-pC%29-RBS32-ccdB-TT_BBa_K1075047.jpg' width='840' height='200'></a>pDawn-RBS32-ccdB-TT can be used to photocontrol the expression of the toxin ccdB. In this system pDawn (BBa_K1075044) can be regulated by blue light, while ccdB (BBa_K1075029) acts as a killswitch. The ccd operon of the F plasmid encodes CcdB, a toxin targeting the essential gyrase of Escherichia coli, and CcdA, the unstable antidote that interacts with CcdB to neutralize its toxicity.<sup><a href=http://parts.igem.org/Part:BBa_K1075047>7</a></sup><a href='https://static.igem.org/mediawiki/2013/0/03/Bonn_AraC-pBAD%28D%29-RBS32-SspB%28Core%28-LOV-ipaA-TT_BBa_K1075019.jpg'><img src='https://static.igem.org/mediawiki/2013/0/03/Bonn_AraC-pBAD%28D%29-RBS32-SspB%28Core%28-LOV-ipaA-TT_BBa_K1075019.jpg' width='840' height='200'></a>The part can be used to regulate the dimerization of the two parts of the split sspB. The binding of ipaA to Vincolin is regulated via the light-sensitive LOV domain. Therefore the two sspB parts only come together when the LOV domain was activated with blue light. <sup><a href=http://parts.igem.org/Part:BBa_K1075019>8</a></sup><a href='https://static.igem.org/mediawiki/2013/d/d0/Bonn_J23105-RBS32-SspB%28Core%29-LOV-ipaA-TT_BBa_K1075018.jpg'><img src='https://static.igem.org/mediawiki/2013/d/d0/Bonn_J23105-RBS32-SspB%28Core%29-LOV-ipaA-TT_BBa_K1075018.jpg' width='840' height='200'></a>The part can be used to regulate the dimerization of the two parts of the split sspB. The binding of ipaA to Vincolin is regulated via the light-sensitive LOV domain. Therefore the two sspB parts only come together when the LOV domain was activated with blue light. <sup><a href=http://parts.igem.org/Part:BBa_K1075018>9</a></sup><a href='https://static.igem.org/mediawiki/2013/5/5a/Bonn_AraC-pBAD%28D%29-RBS32-VinD1-sspBXB-TT_BBa_K1075016.jpg'><img src='https://static.igem.org/mediawiki/2013/5/5a/Bonn_AraC-pBAD%28D%29-RBS32-VinD1-sspBXB-TT_BBa_K1075016.jpg' width='840' height='200'></a>The part can be used to regulate the dimerization of the two parts of the split sspB. As Vincolin bins to ipaA, the sspB dimer is stabilized. We used this part to achieve a light-induced dimerization of the sspB parts.  <sup><a href=http://parts.igem.org/Part:BBa_K1075016>10</a></sup><a href='https://static.igem.org/mediawiki/2013/9/9c/Bonn_J23118-RBS32-VinD1-SspBXB-TT_BBa_K1075015.jpg'><img src='https://static.igem.org/mediawiki/2013/9/9c/Bonn_J23118-RBS32-VinD1-SspBXB-TT_BBa_K1075015.jpg' width='840' height='200'></a>The part can be used to regulate the dimerization of the two parts of the split sspB. As Vincolin bins to ipaA, the sspB dimer is stabilized. We used this part to achieve a light-induced dimerization of the sspB parts. <sup><a href=http://parts.igem.org/Part:BBa_K1075015>11</a></sup><a href='https://static.igem.org/mediawiki/2013/7/78/Bonn_pLac_pBAD2-RBS32-SspB-TT_BBa_K1075006.jpg'><img src='https://static.igem.org/mediawiki/2013/7/78/Bonn_pLac_pBAD2-RBS32-SspB-TT_BBa_K1075006.jpg' width='840' height='200'></a>The sspB protein is an adaptor responsible for delivering ssrA-tagged substrates to the ClpXP protease in order to enhance their degradation. Prefixed is RBS, the binding site for a ribosome and the pLac promoter. It also includes a double terminator 'TT', which interrupts the translation. <sup><a href=http://parts.igem.org/Part:BBa_K1075006>12</a></sup><a href='https://static.igem.org/mediawiki/2013/8/8f/Bonn_E._coli_ssrA%28DAS%2B4%29-TT_BBa_K1075021.jpg'><img src='https://static.igem.org/mediawiki/2013/8/8f/Bonn_E._coli_ssrA%28DAS%2B4%29-TT_BBa_K1075021.jpg' width='840' height='200'></a>The (ec)ssrA(DAS+4) tag can be fused to proteins so that they will be delivered to the ClpXP protease and degraded.<sup><a href=http://parts.igem.org/Part:BBa_K1075021>13</a></sup><a href='https://static.igem.org/mediawiki/2013/6/63/Bonn_AraC-pBad-RBS34-mCherry-ecssrA%28DAS%2B4%29-TT_BBa_K1075025.jpg'><img src='https://static.igem.org/mediawiki/2013/6/63/Bonn_AraC-pBad-RBS34-mCherry-ecssrA%28DAS%2B4%29-TT_BBa_K1075025.jpg' width='840' height='200'></a>As we want to control protein degradation by controlling the function of ecSspB, we tagged the red fluorescent protein mCherry with ecssrA(DAS+4) to measure the degradation rate. The part was designed for proof of principle. Application as a bacterial fotographic film might be possible as well. <sup><a href=http://parts.igem.org/Part:BBa_K1075025>14</a></sup><a href='https://static.igem.org/mediawiki/2013/6/63/Bonn_AraC-pBad-RBS34-mCherry-ecssrA%28DAS%2B4%29-TT_BBa_K1075025.jpg'><img src='https://static.igem.org/mediawiki/2013/6/63/Bonn_AraC-pBad-RBS34-mCherry-ecssrA%28DAS%2B4%29-TT_BBa_K1075025.jpg' width='840' height='200'></a>As we want to control protein degradation by controlling the function of ecSspB, we tagged the red fluorescent protein mCherry with ecssrA(DAS+4) to measure the degradation rate. The part was designed for proof of principle. Application as a bacterial fotographic film might be possible as well. <sup><a href=http://parts.igem.org/Part:BBa_K1075025>15</a></sup><a href='https://static.igem.org/mediawiki/2013/c/cd/Bonn_pJD427%28pC-RBS32-FKBP12-EcsspB%28XB%29-Term-pB-RBS32-EcsspB%28Core%29-FRB-Term-Term%29_BBa_K1075010.jpg'><img src='https://static.igem.org/mediawiki/2013/c/cd/Bonn_pJD427%28pC-RBS32-FKBP12-EcsspB%28XB%29-Term-pB-RBS32-EcsspB%28Core%29-FRB-Term-Term%29_BBa_K1075010.jpg' width='840' height='200'></a>The SspB protein is an adaptor responsible for delivering ssrA-tagged substrates to the ClpXP protease in order to enhance their degradation. To control degradation it is reasonable to control the function of SspB. That is made by splitting it into two parts, each of which cannot induce degradation on its own. To bring both SspB parts together again for inducible degradation, they were combined with a chemical inducible heterodimerisation system: FRB and FKBP12. These two parts interact in the presence of rapamycin <sup><a href=http://parts.igem.org/Part:BBa_K1075010>16</a></sup><a href='https://static.igem.org/mediawiki/2013/d/d6/Bonn_%28pJD427%29-pLac-RBS34-mCherry-ecssrA%28DAS%2B4%29-TT_BBa_K1075027.jpg'><img src='https://static.igem.org/mediawiki/2013/d/d6/Bonn_%28pJD427%29-pLac-RBS34-mCherry-ecssrA%28DAS%2B4%29-TT_BBa_K1075027.jpg' width='840' height='200'></a>As we want to control protein degradation by controlling the function of ecSspB, we tag the red fluorescent protein mCherry with ecssrA(DAS+4) to measure the degradation rate. We control ecSspB by splitting it into two parts each of which cannot induce degradation on its own but regains function by addition of rapamycin. As the co-transformation of the two plasmids didn’t work, we combined them to one.<sup><a href=http://parts.igem.org/Part:BBa_K1075027>17</a></sup><a href='https://static.igem.org/mediawiki/2013/b/bc/Bonn_pJD427-pLac-RBS34-mCherry-TT_BBa_K1075028.jpg'><img src='https://static.igem.org/mediawiki/2013/b/bc/Bonn_pJD427-pLac-RBS34-mCherry-TT_BBa_K1075028.jpg' width='840' height='200'></a>This plasmid is a negative control to the pJD427-pLac2-RBS34-mCherry-ecssrA(DAS+4)-TT plasmid. <sup><a href=http://parts.igem.org/Part:BBa_K1075028>18</a></sup><a href='https://static.igem.org/mediawiki/2013/c/cd/Bonn_pJD427%28pC-RBS32-FKBP12-EcsspB%28XB%29-Term-pB-RBS32-EcsspB%28Core%29-FRB-Term-Term%29_BBa_K1075010.jpg'><img src='https://static.igem.org/mediawiki/2013/c/cd/Bonn_pJD427%28pC-RBS32-FKBP12-EcsspB%28XB%29-Term-pB-RBS32-EcsspB%28Core%29-FRB-Term-Term%29_BBa_K1075010.jpg' width='840' height='200'></a>The Plasmid pJD427 contains the fusion proteins SspB[CORE]-FRB and FKBP12-SspB[XB]: SspB[CORE]-FRB with the weak constitutive promoter proB, FKBP12-SspB[XB] with the strong constitutive promoter proC and a medium-copy p15a origin of replication.  <sup><a href=http://parts.igem.org/Part:BBa_K1075010>19</a></sup><a href='https://static.igem.org/mediawiki/2013/e/e1/Bonn_pLac-RBS32-ccdA-ssrA-TT_BBa_K1075035.jpg'><img src='https://static.igem.org/mediawiki/2013/e/e1/Bonn_pLac-RBS32-ccdA-ssrA-TT_BBa_K1075035.jpg' width='840' height='200'></a>CcdA inhibits the toxin ccdB by binding to it and thus represses cell death. The part contains ccdA with a ssrA tag under the control of the Lac operon. <sup><a href=http://parts.igem.org/Part:BBa_K1075035>20</a></sup><a href='https://static.igem.org/mediawiki/2013/8/81/Bonn_AraC-pBAD%28D%29-RBS32-ccdB-TT_BBa_K1075031.jpg'><img src='https://static.igem.org/mediawiki/2013/8/81/Bonn_AraC-pBAD%28D%29-RBS32-ccdB-TT_BBa_K1075031.jpg' width='840' height='200'></a>The part contains a chemically induced kill-switch. When Arabinose is added to the bacteria, the toxin ccdB is expressed and cell death is initiated. <sup><a href=http://parts.igem.org/Part:BBa_K1075031>20</a></sup>";
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