http://2010.igem.org/wiki/index.php?title=Team:Kyoto/Project/Goal_C&feed=atom&action=historyTeam:Kyoto/Project/Goal C - Revision history2024-03-28T19:26:26ZRevision history for this page on the wikiMediaWiki 1.16.5http://2010.igem.org/wiki/index.php?title=Team:Kyoto/Project/Goal_C&diff=208322&oldid=prevTasuku: /* Goal C: Characterization of the anti-killer gene */2010-10-28T03:55:32Z<p><span class="autocomment">Goal C: Characterization of the anti-killer gene</span></p>
<a href="http://2010.igem.org/wiki/index.php?title=Team:Kyoto/Project/Goal_C&diff=208322&oldid=206714">Show changes</a>Tasukuhttp://2010.igem.org/wiki/index.php?title=Team:Kyoto/Project/Goal_C&diff=206714&oldid=prevY.M.: /* Conclusion */2010-10-28T03:10:38Z<p><span class="autocomment">Conclusion</span></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Conclusion===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Conclusion===</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>1)We made S&Delta;TMD1 by deleting TMD1 from S gene and registed it<partinfo><del class="diffchange diffchange-inline">BBa_K58012</del></partinfo> and lysis cassette&Delta;TMD1(S&Delta;TMD1RRz)<partinfo>BBa_K358014</partinfo> in BioBrick as new parts. </div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>1)We made S&Delta;TMD1 by deleting TMD1 from S gene and registed it<partinfo><ins class="diffchange diffchange-inline">BBa_K358012</ins></partinfo> and lysis cassette&Delta;TMD1(S&Delta;TMD1RRz)<partinfo>BBa_K358014</partinfo> in BioBrick as new parts. </div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>2)By the comparison of the function as killer gene between lysis cassette and lysis cassette&Delta;TMD1, We made sure that TMD1 is essential for fuction of lysis cassette as killer gene.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>2)By the comparison of the function as killer gene between lysis cassette and lysis cassette&Delta;TMD1, We made sure that TMD1 is essential for fuction of lysis cassette as killer gene.</div></td></tr>
</table>Y.M.http://2010.igem.org/wiki/index.php?title=Team:Kyoto/Project/Goal_C&diff=206616&oldid=prevY.M.: /* Conclusion */2010-10-28T03:07:20Z<p><span class="autocomment">Conclusion</span></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Conclusion===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Conclusion===</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>1)We made S&Delta;TMD1 by deleting TMD1 from S gene and registed it and lysis cassette&Delta;TMD1(S&Delta;TMD1RRz) in BioBrick as new parts. </div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>1)We made S&Delta;TMD1 by deleting TMD1 from S gene and registed it<ins class="diffchange diffchange-inline"><partinfo>BBa_K58012</partinfo> </ins>and lysis cassette&Delta;TMD1(S&Delta;TMD1RRz)<ins class="diffchange diffchange-inline"><partinfo>BBa_K358014</partinfo> </ins>in BioBrick as new parts. </div></td></tr>
<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> </div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>2)By the comparison of the function as killer gene between lysis cassette and lysis cassette&Delta;TMD1, We made sure that TMD1 is essential for fuction of lysis cassette as killer gene.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>2)By the comparison of the function as killer gene between lysis cassette and lysis cassette&Delta;TMD1, We made sure that TMD1 is essential for fuction of lysis cassette as killer gene.</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>3)By co-expressing lysis cassette regulated by a lactose promoter, we characterized the function of S&Delta;TMD1 as anti-killer gene. </div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> </div></td></tr>
<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>3)By co-expressing lysis cassette regulated by a lactose promoter, we characterized the function of S&Delta;TMD1 as anti-killer gene.</div></td></tr>
<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div> </div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Reference===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Reference===</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>----</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>----</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div># <html><a name="RefL001"></a></html>Rebecca White, Tram Anh T. Tran, Chelsey A. Dankenbring, John Deaton, and Ry Young.2010.The N-Terminal Transmembrane Domain of _ S Is Required for Holin but Not Antiholin Function_. J. Bacteriol. 192: 725–733 </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div># <html><a name="RefL001"></a></html>Rebecca White, Tram Anh T. Tran, Chelsey A. Dankenbring, John Deaton, and Ry Young.2010.The N-Terminal Transmembrane Domain of _ S Is Required for Holin but Not Antiholin Function_. J. Bacteriol. 192: 725–733 </div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div># <html><a name="RefL002"></a></html>CHUNG-YU CHANG, KIEBANG NAM, AND RY YOUNG1. 1995.2 S Gene Expression and the Timing of Lysis by Bacteriophage λ. J. Bacteriol. 177: 3283–3294</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div># <html><a name="RefL002"></a></html>CHUNG-YU CHANG, KIEBANG NAM, AND RY YOUNG1. 1995.2 S Gene Expression and the Timing of Lysis by Bacteriophage λ. J. Bacteriol. 177: 3283–3294</div></td></tr>
</table>Y.M.http://2010.igem.org/wiki/index.php?title=Team:Kyoto/Project/Goal_C&diff=206474&oldid=prevY.M. at 03:02, 28 October 20102010-10-28T03:02:57Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Conclusion===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Conclusion===</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>1) We made S&Delta;TMD1 by deleting TMD1 from S gene and registed it and lysis cassette&Delta;TMD1(S&Delta;TMD1RRz) in BioBrick as new parts. </div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>1)We made S&Delta;TMD1 by deleting TMD1 from S gene and registed it and lysis cassette&Delta;TMD1(S&Delta;TMD1RRz) in BioBrick as new parts. </div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>2) By the comparison of the function as killer gene between lysis cassette and lysis cassette&Delta;TMD1, We made sure that TMD1 is essential for fuction of lysis cassette as killer gene.</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>2)By the comparison of the function as killer gene between lysis cassette and lysis cassette&Delta;TMD1, We made sure that TMD1 is essential for fuction of lysis cassette as killer gene.</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>3) By co-expressing lysis cassette regulated by a lactose promoter, we characterized the function of S&Delta;TMD1 as anti-killer gene. </div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>3)By co-expressing lysis cassette regulated by a lactose promoter, we characterized the function of S&Delta;TMD1 as anti-killer gene. </div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Reference===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Reference===</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>----</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>----</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div># <html><a name="RefL001"></a></html>Rebecca White, Tram Anh T. Tran, Chelsey A. Dankenbring, John Deaton, and Ry Young.2010.The N-Terminal Transmembrane Domain of _ S Is Required for Holin but Not Antiholin Function_. J. Bacteriol. 192: 725–733 </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div># <html><a name="RefL001"></a></html>Rebecca White, Tram Anh T. Tran, Chelsey A. Dankenbring, John Deaton, and Ry Young.2010.The N-Terminal Transmembrane Domain of _ S Is Required for Holin but Not Antiholin Function_. J. Bacteriol. 192: 725–733 </div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div># <html><a name="RefL002"></a></html>CHUNG-YU CHANG, KIEBANG NAM, AND RY YOUNG1. 1995.2 S Gene Expression and the Timing of Lysis by Bacteriophage λ. J. Bacteriol. 177: 3283–3294</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div># <html><a name="RefL002"></a></html>CHUNG-YU CHANG, KIEBANG NAM, AND RY YOUNG1. 1995.2 S Gene Expression and the Timing of Lysis by Bacteriophage λ. J. Bacteriol. 177: 3283–3294</div></td></tr>
</table>Y.M.http://2010.igem.org/wiki/index.php?title=Team:Kyoto/Project/Goal_C&diff=206401&oldid=prevY.M. at 03:00, 28 October 20102010-10-28T03:00:34Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Then, we checked if S&Delta;TMD1 prevents lysis cassette from causing the cell death. </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Then, we checked if S&Delta;TMD1 prevents lysis cassette from causing the cell death. </div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>To investigate the function of S&DELTA;TMD1, we used the E.coli transformed with both killer and anti-killer genes ( <partinfo>BBa_K358021</partinfo>).</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>To investigate the function of S&DELTA;TMD1, we used the E.coli transformed with both killer and anti-killer genes ( <partinfo>BBa_K358021</partinfo>).</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>Because E.coli will be killed immediately if the killer-gene is constitutively expressed, we constructed the lysis cassette regulated by lac promoter. At first, the E.coli bacteria were grown in the medium without IPTG. After the enough population of E.coli, they were cultured in the one with IPTG to express lysis cassette. After that, we checked the growth rate of E.coli by measuring the absorbance (<del class="diffchange diffchange-inline">A550</del>) of the growth medium. Thus, we investigated the possibility whether E.coli that contains both the killer and anti-killer genes could alive due to the constitutive expression of S&DELTA;TMD1. </div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>Because E.coli will be killed immediately if the killer-gene is constitutively expressed, we constructed the lysis cassette regulated by lac promoter. At first, the E.coli bacteria were grown in the medium without IPTG. After the enough population of E.coli, they were cultured in the one with IPTG to express lysis cassette. After that, we checked the growth rate of E.coli by measuring the absorbance (<ins class="diffchange diffchange-inline">OD 550</ins>) of the growth medium. Thus, we investigated the possibility whether E.coli that contains both the killer and anti-killer genes could alive due to the constitutive expression of S&DELTA;TMD1. </div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Method===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Method===</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>====Mesurement====</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>====Mesurement====</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=====Experiment 1=====</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=====Experiment 1=====</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>We pick up three colonies from each plate, and cultivate them in the supplemented M9 medium for a night, overnight. The overnight cultures were diluted to 0.1~0.11mM in pre-warmed fresh the supplemented M9 medium. 18h later, we measure <del class="diffchange diffchange-inline">A550 </del>of the culture containing 0mM or 1mM IPTG.</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>We pick up three colonies from each plate, and cultivate them in the supplemented M9 medium for a night, overnight. The overnight cultures were diluted to 0.1~0.11mM in pre-warmed fresh the supplemented M9 medium. 18h later, we measure <ins class="diffchange diffchange-inline">OD 550 </ins>of the culture containing 0mM or 1mM IPTG.</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=====Experiment 2=====</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=====Experiment 2=====</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>We pick up three colonies from each plate, and cultivate them in the supplemented M9 medium ~24-30 h (overnight).. The overnight cultures were diluted to 0.1~0.11 in pre-warmed fresh the supplemented M9 medium. we measure <del class="diffchange diffchange-inline">A550 </del>of the culture containing various IPTG at about 100 minute interval.</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>We pick up three colonies from each plate, and cultivate them in the supplemented M9 medium ~24-30 h (overnight).. The overnight cultures were diluted to 0.1~0.11 in pre-warmed fresh the supplemented M9 medium. we measure <ins class="diffchange diffchange-inline">OD 550 </ins>of the culture containing various IPTG at about 100 minute interval.</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Result===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Result===</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>[[image:GoalC fig1.png|440px|Fig.1|left|thumb|Fig.2.the result of function check of S&Delta;TMD1.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>[[image:GoalC fig1.png|440px|Fig.1|left|thumb|Fig.2.the result of function check of S&Delta;TMD1.</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>We measure <del class="diffchange diffchange-inline">A </del>550 changing IPTG concentration, 0, 0.02, 0.03, 0.05, 0.3, 0.5, 0.7, 1.0 (mM). ]]</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>We measure <ins class="diffchange diffchange-inline">OD </ins>550 changing IPTG concentration, 0, 0.02, 0.03, 0.05, 0.3, 0.5, 0.7, 1.0 (mM). ]]</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>[[image:GoalC fig2-3-2.png|440px|right|thumb|Fig.3.Comparison between lacL and LB2-1.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>[[image:GoalC fig2-3-2.png|440px|right|thumb|Fig.3.Comparison between lacL and LB2-1.</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>So, we assume that the first reason is negative.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>So, we assume that the first reason is negative.</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=====Reason2:S&Delta;TMD1 is weaker anti-killer gene than we expected.=====</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=====Reason2:S&Delta;TMD1 is weaker anti-killer gene than we expected.=====</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>As anti-killer gene against lysis cassette, there are two genes, S107 and S&Delta;TMD1([[Team:Kyoto/LearnMore#Lysis Cassette|Learn more]])<del class="diffchange diffchange-inline">. </del>. </div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>As anti-killer gene against lysis cassette, there are two genes, S107 and S&Delta;TMD1([[Team:Kyoto/LearnMore#Lysis Cassette|Learn more]]).</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><del class="diffchange diffchange-inline">. </del>S107 is the anti-killer gene. Because it has essential TMD1 of S for function of this killer gene, the anti-killer function of this gene is weak compared with S&Delta;TMD1, which does not have TMD1([[Team:Kyoto/LearnMore#Lysis Cassette|Learn more]]).</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>S107 is the anti-killer gene. Because it has essential TMD1 of S for function of this killer gene, the anti-killer function of this gene is weak compared with S&Delta;TMD1, which does not have TMD1([[Team:Kyoto/LearnMore#Lysis Cassette|Learn more]]).</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Thus, we expected that S&Delta;TMD1 is a strong anti-killer gene. However, from the result of experiment 2, we could not observe the function of S&Delta;TMD1 as a strong anti-killer gene under the tested condition.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Thus, we expected that S&Delta;TMD1 is a strong anti-killer gene. However, from the result of experiment 2, we could not observe the function of S&Delta;TMD1 as a strong anti-killer gene under the tested condition.</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=====Reason3: Expression balance between the anti-killer and killer genes was not optimized properly.=====</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=====Reason3: Expression balance between the anti-killer and killer genes was not optimized properly.=====</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>We assume that our experimental results could be due to the 3rd reason (Reason 3) mentioned above, although we can not rule out the possibility of the second reason. However, it has been reported <sup>[[#RefL001|[1]]]</sup> that S&Delta;TMD1 inhibits lysis cassette effectively. So, it is reasonable to assume that S&Delta;TMD1 exhibits enough activity to inhibit lysis cassette. That is why we think that the Reason 3 is most likely the answer. To confirm this possibility, we changed the promoter of S&Delta;TMD1 to stronger one. The experiment to check the function of S&Delta;TMD1 using the strong promoter is currently underway. Surely, this result will help us to understand why S&Delta;TMD1 could not function as we expected and to develop distinct cell-death controllable devices for the further studies. </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>We assume that our experimental results could be due to the 3rd reason (Reason 3) mentioned above, although we can not rule out the possibility of the second reason. However, it has been reported <sup>[[#RefL001|[1]]]</sup> that S&Delta;TMD1 inhibits lysis cassette effectively. So, it is reasonable to assume that S&Delta;TMD1 exhibits enough activity to inhibit lysis cassette. That is why we think that the Reason 3 is most likely the answer. To confirm this possibility, we changed the promoter of S&Delta;TMD1 to stronger one. The experiment to check the function of S&Delta;TMD1 using the strong promoter is currently underway. Surely, this result will help us to understand why S&Delta;TMD1 could not function as we expected and to develop distinct cell-death controllable devices for the further studies. </div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><del class="diffchange diffchange-inline">=</del>===Conclusion<del class="diffchange diffchange-inline">=</del>===</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>===Conclusion===</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>1) We made S&Delta;TMD1 by deleting TMD1 from S gene and registed it and lysis cassette&Delta;TMD1(S&Delta;TMD1RRz) in BioBrick as new parts. </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>1) We made S&Delta;TMD1 by deleting TMD1 from S gene and registed it and lysis cassette&Delta;TMD1(S&Delta;TMD1RRz) in BioBrick as new parts. </div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>2) By the comparison of the function as killer gene between lysis cassette and lysis cassette&Delta;TMD1, We made sure that TMD1 is essential for fuction of lysis cassette as killer gene.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>2) By the comparison of the function as killer gene between lysis cassette and lysis cassette&Delta;TMD1, We made sure that TMD1 is essential for fuction of lysis cassette as killer gene.</div></td></tr>
</table>Y.M.http://2010.igem.org/wiki/index.php?title=Team:Kyoto/Project/Goal_C&diff=206160&oldid=prevY.M. at 02:52, 28 October 20102010-10-28T02:52:55Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>{{clear}}</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>{{clear}}</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>*<partinfo>BBa_K358021</partinfo>:Anti-killergene is induced constitutively and killergene is regulated by lactose promoter.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>*<partinfo>BBa_K358021</partinfo>:Anti-killergene is induced constitutively and killergene is regulated by lactose promoter.</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>It was used in experiment 2</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>It was used in experiment 2<ins class="diffchange diffchange-inline">.</ins></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>*<partinfo>BBa_K358022</partinfo>:lacP + S&DELTA;TMD1RRz, TMD1 is deleted from lysis cassette[SRRz].</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>*<partinfo>BBa_K358022</partinfo>:lacP + S&DELTA;TMD1RRz, TMD1 is deleted from lysis cassette[SRRz].</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>It was used in experiment 1 </div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>It was used in experiment 1<ins class="diffchange diffchange-inline">. </ins></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>*<partinfo>BBa_K358024</partinfo>:<del class="diffchange diffchange-inline"> This </del>part is the same to BBa_K358021, except for its constitutive promoter[BBa_J23101]. This part also induces constitutively S&DELTA;TMD1 gene, the anti-killer gene, and lambda lysis cassette[SRRz], the killer gene, is regulated by lacP.</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>*<partinfo>BBa_K358024</partinfo>: <ins class="diffchange diffchange-inline">This </ins>part is the same to BBa_K358021, except for its constitutive promoter[BBa_J23101]. This part also induces constitutively S&DELTA;TMD1 gene, the anti-killer gene, and lambda lysis cassette[SRRz], the killer gene, is regulated by lacP.</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>It was used in the additional experiment, which is not finished.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>It was used in the additional experiment, which is not finished.</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>*<partinfo>BBa_K358019</partinfo>: Lysis cassette regulated by lacP, Lysis cassette[SRRz] is regulated by lactose promoter. </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>*<partinfo>BBa_K358019</partinfo>: Lysis cassette regulated by lacP, Lysis cassette[SRRz] is regulated by lactose promoter. </div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Discussion===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Discussion===</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>====TMD1 of S gene is essential for lysis cassette====</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>====TMD1 of S gene is essential for lysis cassette====</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>The result of experiment 1 supports [[#RefL001|Ref.1].</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>The result of experiment 1 supports [[#RefL001|Ref.1<ins class="diffchange diffchange-inline">]</ins>].</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>That is, we were able to make sure that TMD1 of S gene is essential for lysis cassette as killer gene.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>That is, we were able to make sure that TMD1 of S gene is essential for lysis cassette as killer gene.</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>So, we assume that the first reason is negative.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>So, we assume that the first reason is negative.</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=====Reason2:S&Delta;TMD1 is weaker anti-killer gene than we expected.=====</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=====Reason2:S&Delta;TMD1 is weaker anti-killer gene than we expected.=====</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>As anti-killer gene against lysis cassette, there are two genes, S107 and S&Delta;TMD1([[Team:Kyoto/LearnMore#Lysis Cassette|Learn more]]).</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>As anti-killer gene against lysis cassette, there are two genes, S107 and S&Delta;TMD1([[Team:Kyoto/LearnMore#Lysis Cassette|Learn more]])<ins class="diffchange diffchange-inline">. </ins>. </div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>S107 is the anti-killer gene. Because it has essential TMD1 of S for function of this killer gene, the anti-killer function of this gene is weak compared with S&Delta;TMD1, which does not have TMD1([[Team:Kyoto/LearnMore#Lysis Cassette|Learn more]]).</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins class="diffchange diffchange-inline">. </ins>S107 is the anti-killer gene. Because it has essential TMD1 of S for function of this killer gene, the anti-killer function of this gene is weak compared with S&Delta;TMD1, which does not have TMD1([[Team:Kyoto/LearnMore#Lysis Cassette|Learn more]]).</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Thus, we expected that S&Delta;TMD1 is a strong anti-killer gene. However, from the result of experiment 2, we could not observe the function of S&Delta;TMD1 as a strong anti-killer gene under the tested condition.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Thus, we expected that S&Delta;TMD1 is a strong anti-killer gene. However, from the result of experiment 2, we could not observe the function of S&Delta;TMD1 as a strong anti-killer gene under the tested condition.</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=====Reason3: Expression balance between the anti-killer and killer genes was not optimized properly.=====</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=====Reason3: Expression balance between the anti-killer and killer genes was not optimized properly.=====</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>====Conclusion====</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>====Conclusion====</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>1) We made S&Delta;TMD1 by deleting TMD1 from <del class="diffchange diffchange-inline">Sgene </del></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>1) We made S&Delta;TMD1 by deleting TMD1 from <ins class="diffchange diffchange-inline">S gene and registed it and lysis cassette&Delta;TMD1(S&Delta;TMD1RRz) in BioBrick as new parts. </ins></div></td></tr>
<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins class="diffchange diffchange-inline">2) By the comparison of the function as killer gene between lysis cassette and lysis cassette&Delta;TMD1, We made sure that TMD1 is essential for fuction of lysis cassette as killer gene.</ins></div></td></tr>
<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins class="diffchange diffchange-inline">3) By co-expressing lysis cassette regulated by a lactose promoter, we characterized the function of S&Delta;TMD1 as anti-killer gene. </ins></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Reference===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Reference===</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>----</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>----</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div># <html><a name="RefL001"></a></html>Rebecca White, Tram Anh T. Tran, Chelsey A. Dankenbring, John Deaton, and Ry Young.2010.The N-Terminal Transmembrane Domain of _ S Is Required for Holin but Not Antiholin Function_. J. Bacteriol. 192: 725–733 </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div># <html><a name="RefL001"></a></html>Rebecca White, Tram Anh T. Tran, Chelsey A. Dankenbring, John Deaton, and Ry Young.2010.The N-Terminal Transmembrane Domain of _ S Is Required for Holin but Not Antiholin Function_. J. Bacteriol. 192: 725–733 </div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div># <html><a name="RefL002"></a></html>CHUNG-YU CHANG, KIEBANG NAM, AND RY YOUNG1. 1995.2 S Gene Expression and the Timing of Lysis by Bacteriophage λ. J. Bacteriol. 177: 3283–3294</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div># <html><a name="RefL002"></a></html>CHUNG-YU CHANG, KIEBANG NAM, AND RY YOUNG1. 1995.2 S Gene Expression and the Timing of Lysis by Bacteriophage λ. J. Bacteriol. 177: 3283–3294</div></td></tr>
</table>Y.M.http://2010.igem.org/wiki/index.php?title=Team:Kyoto/Project/Goal_C&diff=205682&oldid=prevY.M. at 02:36, 28 October 20102010-10-28T02:36:22Z<p></p>
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<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;">{{:Team:Kyoto/Header}}</ins></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==Goal C: Characterization of the anti-killer gene==</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==Goal C: Characterization of the anti-killer gene==</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Introduction===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Introduction===</div></td></tr>
</table>Y.M.http://2010.igem.org/wiki/index.php?title=Team:Kyoto/Project/Goal_C&diff=205596&oldid=prevY.M. at 02:32, 28 October 20102010-10-28T02:32:11Z<p></p>
<table style="background-color: white; color:black;">
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>So, we assume that the first reason is negative.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>So, we assume that the first reason is negative.</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=====Reason2:S&Delta;TMD1 is weaker anti-killer gene than we expected.=====</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=====Reason2:S&Delta;TMD1 is weaker anti-killer gene than we expected.=====</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>As anti-killer gene against lysis cassette, there are two genes, S107 and S&Delta;TMD1([[Team:Kyoto/LearnMore#Lysis Cassette|Learn more]])<del class="diffchange diffchange-inline">. </del>. </div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>As anti-killer gene against lysis cassette, there are two genes, S107 and S&Delta;TMD1([[Team:Kyoto/LearnMore#Lysis Cassette|Learn more]]).</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><del class="diffchange diffchange-inline">. </del>S107 is the anti-killer gene. Because it has essential TMD1 of S for function of this killer gene, the anti-killer function of this gene is weak compared with S&Delta;TMD1, which does not have TMD1([[Team:Kyoto/LearnMore#Lysis Cassette|Learn more]]).</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>S107 is the anti-killer gene. Because it has essential TMD1 of S for function of this killer gene, the anti-killer function of this gene is weak compared with S&Delta;TMD1, which does not have TMD1([[Team:Kyoto/LearnMore#Lysis Cassette|Learn more]]).</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Thus, we expected that S&Delta;TMD1 is a strong anti-killer gene. However, from the result of experiment 2, we could not observe the function of S&Delta;TMD1 as a strong anti-killer gene under the tested condition.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Thus, we expected that S&Delta;TMD1 is a strong anti-killer gene. However, from the result of experiment 2, we could not observe the function of S&Delta;TMD1 as a strong anti-killer gene under the tested condition.</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=====Reason3: Expression balance between the anti-killer and killer genes was not optimized properly.=====</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=====Reason3: Expression balance between the anti-killer and killer genes was not optimized properly.=====</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>====Conclusion====</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>====Conclusion====</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>1) We made S&Delta;TMD1 by deleting TMD1 <del class="diffchange diffchange-inline">of </del>Sgene </div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>1) We made S&Delta;TMD1 by deleting TMD1 <ins class="diffchange diffchange-inline">from </ins>Sgene </div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Reference===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Reference===</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>----</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>----</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div># <html><a name="RefL001"></a></html>Rebecca White, Tram Anh T. Tran, Chelsey A. Dankenbring, John Deaton, and Ry Young.2010.The N-Terminal Transmembrane Domain of _ S Is Required for Holin but Not Antiholin Function_. J. Bacteriol. 192: 725–733 </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div># <html><a name="RefL001"></a></html>Rebecca White, Tram Anh T. Tran, Chelsey A. Dankenbring, John Deaton, and Ry Young.2010.The N-Terminal Transmembrane Domain of _ S Is Required for Holin but Not Antiholin Function_. J. Bacteriol. 192: 725–733 </div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div># <html><a name="RefL002"></a></html>CHUNG-YU CHANG, KIEBANG NAM, AND RY YOUNG1. 1995.2 S Gene Expression and the Timing of Lysis by Bacteriophage λ. J. Bacteriol. 177: 3283–3294</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div># <html><a name="RefL002"></a></html>CHUNG-YU CHANG, KIEBANG NAM, AND RY YOUNG1. 1995.2 S Gene Expression and the Timing of Lysis by Bacteriophage λ. J. Bacteriol. 177: 3283–3294</div></td></tr>
</table>Y.M.http://2010.igem.org/wiki/index.php?title=Team:Kyoto/Project/Goal_C&diff=205567&oldid=prevY.M. at 02:30, 28 October 20102010-10-28T02:30:14Z<p></p>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><del style="color: red; font-weight: bold; text-decoration: none;">{{:Team:Kyoto/Header}}</del></div></td><td colspan="2"> </td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==Goal C: Characterization of the anti-killer gene==</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>==Goal C: Characterization of the anti-killer gene==</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Introduction===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Introduction===</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>As long as we know, there are two anti-killer <del class="diffchange diffchange-inline">gene against </del>the killer gene<del class="diffchange diffchange-inline">,</del>lysis cassette, S107 and S&Delta;TMD1.</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>As long as we know, there are two anti-killer <ins class="diffchange diffchange-inline">genes against </ins>the killer gene<ins class="diffchange diffchange-inline">(</ins>lysis cassette<ins class="diffchange diffchange-inline">)</ins>, S107 and S&Delta;TMD1.</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>S107 is expressed by S gene encoded by lysis cassette and S&Delta;TMD1 is an S allele with the transmembrane domein 1(TMD1) deleted.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>S107 is expressed by S gene encoded by lysis cassette and S&Delta;TMD1 is an S allele with the transmembrane domein 1(TMD1) deleted.</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Because TMD1 of S gene is essential for the function of lysis cassette as killer gene, S&Delta;TMD1 inhibits lysis cassette more strongly than S107.([[Team:Kyoto/LearnMore#Lysis Cassette|Learn more]])</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Because TMD1 of S gene is essential for the function of lysis cassette as killer gene, S&Delta;TMD1 inhibits lysis cassette more strongly than S107.([[Team:Kyoto/LearnMore#Lysis Cassette|Learn more]])</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Then, we checked if S&Delta;TMD1 prevents lysis cassette from causing the cell death. </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>Then, we checked if S&Delta;TMD1 prevents lysis cassette from causing the cell death. </div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>To <del class="diffchange diffchange-inline">research this </del>function of S&<del class="diffchange diffchange-inline">Delta</del>;TMD1, we used the E.coli transformed with both <del class="diffchange diffchange-inline">these </del>genes( <partinfo>BBa_K358021</partinfo>).</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>To <ins class="diffchange diffchange-inline">investigate the </ins>function of S&<ins class="diffchange diffchange-inline">DELTA</ins>;TMD1, we used the E.coli transformed with both <ins class="diffchange diffchange-inline">killer and anti-killer </ins>genes ( <partinfo>BBa_K358021</partinfo>).</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>Because E.coli <del class="diffchange diffchange-inline">is </del>immediately <del class="diffchange diffchange-inline">dead </del>if killer-gene is constitutively expressed, we <del class="diffchange diffchange-inline">made </del>lysis cassette regulated by lac promoter. <del class="diffchange diffchange-inline">The </del>E.coli bacteria were grown in the medium without IPTG<del class="diffchange diffchange-inline">, and after there were </del>enough E.coli, they were cultured in the one with IPTG to express lysis cassette. After that, we checked<del class="diffchange diffchange-inline">, </del>by measuring the absorbance (A550), <del class="diffchange diffchange-inline">if </del>E.coli <del class="diffchange diffchange-inline">were </del>alive due to S&<del class="diffchange diffchange-inline">Delta</del>;TMD1 <del class="diffchange diffchange-inline">expressed constitutively</del>.</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>Because E.coli <ins class="diffchange diffchange-inline">will be killed </ins>immediately if <ins class="diffchange diffchange-inline">the </ins>killer-gene is constitutively expressed, we <ins class="diffchange diffchange-inline">constructed the </ins>lysis cassette regulated by lac promoter. <ins class="diffchange diffchange-inline">At first, the </ins>E.coli bacteria were grown in the medium without IPTG<ins class="diffchange diffchange-inline">. After the </ins>enough <ins class="diffchange diffchange-inline">population of </ins>E.coli, they were cultured in the one with IPTG to express lysis cassette. After that, we checked <ins class="diffchange diffchange-inline">the growth rate of E.coli </ins>by measuring the absorbance (A550) <ins class="diffchange diffchange-inline">of the growth medium. Thus</ins>, <ins class="diffchange diffchange-inline">we investigated the possibility whether </ins>E.coli <ins class="diffchange diffchange-inline">that contains both the killer and anti-killer genes could </ins>alive due to <ins class="diffchange diffchange-inline">the constitutive expression of </ins>S&<ins class="diffchange diffchange-inline">DELTA</ins>;TMD1. </div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Method===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Method===</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>====Bacterial strains====</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>====Bacterial strains====</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>We used four types of E. coli<del class="diffchange diffchange-inline">, </del>E. coli KRX transformed with <partinfo>BBa_K358021</partinfo>, KRX transformed with <partinfo>BBa_K358022</partinfo><del class="diffchange diffchange-inline"> KRX </del>transformed with <partinfo>BBa_K358024</partinfo> and <del class="diffchange diffchange-inline">KPX </del>transfomed with <partinfo>BBa_K358019</partinfo>.</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>We used four types of E. coli <ins class="diffchange diffchange-inline">as follows: (1) </ins>E. coli KRX transformed with <partinfo>BBa_K358021</partinfo>, <ins class="diffchange diffchange-inline">(2)</ins>KRX transformed with <partinfo>BBa_K358022</partinfo><ins class="diffchange diffchange-inline">, (3)KRX </ins>transformed with <partinfo>BBa_K358024</partinfo> and <ins class="diffchange diffchange-inline">(4)KRX </ins>transfomed with <partinfo>BBa_K358019</partinfo>.</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>After this, we call each E.coli to abbreviate in order of name, LB2-1, lacLΔ, LB2-2 and lacL.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>After this, we call each E.coli to abbreviate in order of name, LB2-1, lacLΔ, LB2-2 and lacL.</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>*<partinfo>BBa_K358021</partinfo>:Anti-killergene is induced constitutively and killergene is regulated by lactose promoter.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>*<partinfo>BBa_K358021</partinfo>:Anti-killergene is induced constitutively and killergene is regulated by lactose promoter.</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>It was used in experiment 2</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>It was used in experiment 2</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>*<partinfo>BBa_K358022</partinfo>:lacP + <del class="diffchange diffchange-inline">SΔTMD1RRz</del>, TMD1 is deleted from lysis cassette[SRRz].</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>*<partinfo>BBa_K358022</partinfo>:lacP + <ins class="diffchange diffchange-inline">S&DELTA;TMD1RRz</ins>, TMD1 is deleted from lysis cassette[SRRz].</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>It was used in experiment 1 </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>It was used in experiment 1 </div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>*<partinfo>BBa_K358024</partinfo>: This part is the same to BBa_K358021, except for its constitutive promoter[BBa_J23101]. This part also induces constitutively <del class="diffchange diffchange-inline">SΔTMD1 </del>gene, the anti-killer gene, and lambda lysis cassette[SRRz], the killer gene, is regulated by lacP.</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>*<partinfo>BBa_K358024</partinfo>: This part is the same to BBa_K358021, except for its constitutive promoter[BBa_J23101]. This part also induces constitutively <ins class="diffchange diffchange-inline">S&DELTA;TMD1 </ins>gene, the anti-killer gene, and lambda lysis cassette[SRRz], the killer gene, is regulated by lacP.</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>It was used in the additional experiment, which is not finished.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>It was used in the additional experiment, which is not finished.</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>*<partinfo>BBa_K358019</partinfo>: Lysis cassette regulated by lacP, Lysis cassette[SRRz] is regulated by lactose promoter. </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>*<partinfo>BBa_K358019</partinfo>: Lysis cassette regulated by lacP, Lysis cassette[SRRz] is regulated by lactose promoter. </div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=====Experiment 2=====</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=====Experiment 2=====</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>We pick up three colonies from each plate, and cultivate them in the supplemented M9 medium overnight<del class="diffchange diffchange-inline">, about 16 hours</del>. The overnight cultures were diluted to 0.1~0.11 in pre-warmed fresh the supplemented M9 medium. we measure A550 of the culture containing various IPTG at about 100 minute interval.</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>We pick up three colonies from each plate, and cultivate them in the supplemented M9 medium <ins class="diffchange diffchange-inline"> ~24-30 h (</ins>overnight<ins class="diffchange diffchange-inline">).</ins>. The overnight cultures were diluted to 0.1~0.11 in pre-warmed fresh the supplemented M9 medium. we measure A550 of the culture containing various IPTG at about 100 minute interval.</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Result===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Result===</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>*less S&Delta;TMD1 proteins or more lysis cassette proteins was expressed than we expected.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>*less S&Delta;TMD1 proteins or more lysis cassette proteins was expressed than we expected.</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>=====Reason1:S&Delta;TMD1 in the plasmid did not <del class="diffchange diffchange-inline">worked </del>correctly.=====</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>=====Reason1:S&Delta;TMD1 in the plasmid did not <ins class="diffchange diffchange-inline">work </ins>correctly.=====</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>To <del class="diffchange diffchange-inline">make sure the </del>reason, we sequenced BBa_K358021 we used. </div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>To <ins class="diffchange diffchange-inline">investigate this </ins>reason, we sequenced BBa_K358021 we used. </div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><del class="diffchange diffchange-inline">It’s </del>result showed that this construct is inserted into the plasmid correctly, which demonstrated that S&Delta;TMD1 worked correctly.</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins class="diffchange diffchange-inline">The </ins>result showed that this construct is inserted into the plasmid correctly, which demonstrated that S&Delta;TMD1 worked correctly.</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>So, first reason <del class="diffchange diffchange-inline">was not right</del>.</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>So, <ins class="diffchange diffchange-inline">we assume that the </ins>first reason <ins class="diffchange diffchange-inline">is negative</ins>.</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=====Reason2:S&Delta;TMD1 is weaker anti-killer gene than we expected.=====</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>=====Reason2:S&Delta;TMD1 is weaker anti-killer gene than we expected.=====</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>As anti-killer gene against lysis cassette, there are two genes, S107 and S&Delta;TMD1([[Team:Kyoto/LearnMore#Lysis Cassette|Learn more]]).</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>As anti-killer gene against lysis cassette, there are two genes, S107 and S&Delta;TMD1([[Team:Kyoto/LearnMore#Lysis Cassette|Learn more]])<ins class="diffchange diffchange-inline">. </ins>. </div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>S107 is anti-killer gene<del class="diffchange diffchange-inline">, but because </del>it has essential TMD1 of S for function of this killer gene, <del class="diffchange diffchange-inline">it’s function as </del>anti-killer gene is weak compared with S&Delta;TMD1, which does not have TMD1 <del class="diffchange diffchange-inline">of </del>([[Team:Kyoto/LearnMore#Lysis Cassette|Learn more]]).</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins class="diffchange diffchange-inline">. </ins>S107 is <ins class="diffchange diffchange-inline">the </ins>anti-killer gene<ins class="diffchange diffchange-inline">. Because </ins>it has essential TMD1 of S for function of this killer gene, <ins class="diffchange diffchange-inline">the </ins>anti-killer <ins class="diffchange diffchange-inline">function of this </ins>gene is weak compared with S&Delta;TMD1, which does not have TMD1([[Team:Kyoto/LearnMore#Lysis Cassette|Learn more]]).</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><del class="diffchange diffchange-inline">So</del>, we <del class="diffchange diffchange-inline">thought </del>that S&Delta;TMD1 is a strong anti-killer gene, <del class="diffchange diffchange-inline">but </del>from result of experiment <del class="diffchange diffchange-inline">1 </del>we <del class="diffchange diffchange-inline">doubted </del>S&Delta;TMD1 <del class="diffchange diffchange-inline">was </del>a strong anti-killer gene<del class="diffchange diffchange-inline">. However, I could not make sure </del>the <del class="diffchange diffchange-inline">reason</del>.</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins class="diffchange diffchange-inline">Thus</ins>, we <ins class="diffchange diffchange-inline">expected </ins>that S&Delta;TMD1 is a strong anti-killer gene<ins class="diffchange diffchange-inline">. However</ins>, from <ins class="diffchange diffchange-inline">the </ins>result of experiment <ins class="diffchange diffchange-inline">2, </ins>we <ins class="diffchange diffchange-inline">could not observe the function of </ins>S&Delta;TMD1 <ins class="diffchange diffchange-inline">as </ins>a strong anti-killer gene <ins class="diffchange diffchange-inline">under </ins>the <ins class="diffchange diffchange-inline">tested condition</ins>.</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> </div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>=====Reason3: <ins class="diffchange diffchange-inline">Expression balance between the anti-killer and killer genes </ins>was <ins class="diffchange diffchange-inline">not optimized properly</ins>.=====</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>=====Reason3: <del class="diffchange diffchange-inline">less S&Delta;TMD1 proteins or more lysis cassette proteins </del>was <del class="diffchange diffchange-inline">expressed than we expected</del>.=====</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>In order to <ins class="diffchange diffchange-inline">investigate the possibility of </ins>this reason, we <ins class="diffchange diffchange-inline">analyzed </ins>the expression level of <ins class="diffchange diffchange-inline">the </ins>killer gene, lysis cassette and S&Delta;TMD1. </div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>In order to <del class="diffchange diffchange-inline">make sure </del>this reason, we <del class="diffchange diffchange-inline">researched </del>the expression level of killer gene, lysis cassette and S&Delta;TMD1. </div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>As a report <sup>[[#RefL002|[2]]]</sup> says, it is necessary for the inhibition of lysis cassette that there are at least <ins class="diffchange diffchange-inline">~</ins>4.<ins class="diffchange diffchange-inline">7 </ins>times more S107, anti-killer gene than lysis cassette proteins. </div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>As a report <sup>[[#RefL002|[2]]]</sup> says, it is necessary for the inhibition of lysis cassette that there are at least 4.<del class="diffchange diffchange-inline">73 </del>times more S107, anti-killer gene than lysis cassette proteins. </div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>So, Inhibition of this killer gene needs at least <ins class="diffchange diffchange-inline">~</ins>4.<ins class="diffchange diffchange-inline">7 </ins>times more <ins class="diffchange diffchange-inline">expression of the </ins>anti-killer gene than <ins class="diffchange diffchange-inline">that of the </ins>killer gene. <ins class="diffchange diffchange-inline">Thus</ins>, we calculated the ratio of <ins class="diffchange diffchange-inline">the expression levels of the </ins>killer<ins class="diffchange diffchange-inline">- </ins>and anti-killer<ins class="diffchange diffchange-inline">- </ins>gene<ins class="diffchange diffchange-inline">. The calculation was done under the assumptions that </ins>S gene, which lysis cassette includes, produces <ins class="diffchange diffchange-inline">S105 and S107 at a </ins>2.5:1 <ins class="diffchange diffchange-inline">ratio</ins>([[Team:Kyoto/LearnMore#Lysis Cassette|Learn more]]), <ins class="diffchange diffchange-inline">and that the </ins>strength <ins class="diffchange diffchange-inline">of function </ins>of S&<ins class="diffchange diffchange-inline">Delta;TMD1 as anti-killer gene </ins>is the same <ins class="diffchange diffchange-inline">as that of S107</ins>. By this <ins class="diffchange diffchange-inline">calculation</ins>, we knew that in LB-1, the anti-killer gene couldn't inhibit killer gene in the medium with more than 0.<ins class="diffchange diffchange-inline">1246 mM </ins>IPTG. So, we <ins class="diffchange diffchange-inline">assume </ins>that S&Delta;TMD1 was not expressed adequately <ins class="diffchange diffchange-inline">to </ins>inhibit lysis cassette completly. </div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>So, Inhibition of this killer gene needs at least 4.<del class="diffchange diffchange-inline">73 </del>times more anti-killer gene than killer gene. <del class="diffchange diffchange-inline">Then</del>, we calculated the ratio of killer <del class="diffchange diffchange-inline">gene </del>and anti-killer gene<del class="diffchange diffchange-inline">, assuming that </del>S gene, which lysis cassette includes, produces 2.5:1 <del class="diffchange diffchange-inline">S107 and S105</del>([[Team:Kyoto/LearnMore#Lysis Cassette|Learn more]]), strength of S&<del class="diffchange diffchange-inline">DeltaTMD1 and S107 </del>is the same. By this <del class="diffchange diffchange-inline">calcuration</del>, we knew that in LB-1, the anti-killer gene couldn't inhibit killer gene in the medium with more than 0.<del class="diffchange diffchange-inline">1246mM </del>IPTG. So, we <del class="diffchange diffchange-inline">insist </del>that <del class="diffchange diffchange-inline">it is because </del>S&Delta;TMD1 was not expressed <del class="diffchange diffchange-inline"> </del>adequately <del class="diffchange diffchange-inline">that S&Delta;TMD1 cannot </del>inhibit lysis cassette completly.</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>We <ins class="diffchange diffchange-inline">assume </ins>that <ins class="diffchange diffchange-inline">our experimental results could be due to </ins>the 3rd reason <ins class="diffchange diffchange-inline">(Reason 3) mentioned above</ins>, <ins class="diffchange diffchange-inline">although </ins>we <ins class="diffchange diffchange-inline">can </ins>not <ins class="diffchange diffchange-inline">rule out the possibility of the second reason</ins>. However, <ins class="diffchange diffchange-inline">it has been reported </ins><sup>[[#RefL001|[1]]]</sup> that S&Delta;TMD1 inhibits lysis cassette <ins class="diffchange diffchange-inline">effectively</ins>. So, it is <ins class="diffchange diffchange-inline">reasonable to assume </ins>that S&Delta;TMD1 <ins class="diffchange diffchange-inline">exhibits </ins>enough activity to inhibit lysis cassette. That is why <ins class="diffchange diffchange-inline">we think that </ins>the <ins class="diffchange diffchange-inline">Reason 3 </ins>is <ins class="diffchange diffchange-inline">most likely the answer</ins>. <ins class="diffchange diffchange-inline">To confirm </ins>this <ins class="diffchange diffchange-inline">possibility</ins>, we changed the promoter of S&Delta;TMD1 to stronger one<ins class="diffchange diffchange-inline">. The experiment to check </ins>the function of S&Delta;TMD1 <ins class="diffchange diffchange-inline">using the strong promoter is currently underway</ins>. Surely, this result will help us <ins class="diffchange diffchange-inline">to understand </ins>why S&Delta;TMD1 <ins class="diffchange diffchange-inline">could not </ins>function as we expected <ins class="diffchange diffchange-inline">and to develop distinct cell-death controllable devices for the further studies</ins>. </div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> </div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><del class="diffchange diffchange-inline">=====Conclusion=====</del></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>We <del class="diffchange diffchange-inline">insist </del>that the 3rd reason <del class="diffchange diffchange-inline">is right. Certainly</del>, we <del class="diffchange diffchange-inline">have </del>not <del class="diffchange diffchange-inline">made sure that 2nd Reason is not right</del>. However, <del class="diffchange diffchange-inline">a report</del><sup>[[#RefL001|[1]]]</sup> <del class="diffchange diffchange-inline">says </del>that S&Delta;TMD1 inhibits lysis cassette. So, it is <del class="diffchange diffchange-inline">natural </del>that <del class="diffchange diffchange-inline">we think </del>S&Delta;TMD1 <del class="diffchange diffchange-inline">have </del>enough activity to inhibit lysis cassette. </div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>That is why the <del class="diffchange diffchange-inline">3rd reason </del>is <del class="diffchange diffchange-inline">right</del>.</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div> </div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><del class="diffchange diffchange-inline">Then, in order to resolve </del>this <del class="diffchange diffchange-inline">problem</del>, we changed the promoter of S&Delta;TMD1 to stronger one<del class="diffchange diffchange-inline">, and now, are checking </del>the function of S&Delta;TMD1. Surely, this result will help us <del class="diffchange diffchange-inline">discuss </del>why S&Delta;TMD1 <del class="diffchange diffchange-inline">couldn’t </del>function as we expected.</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div></div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><del class="diffchange diffchange-inline">You must expect it!!</del></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"></td></tr>
<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;">====Conclusion====</ins></div></td></tr>
<tr><td colspan="2"> </td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><ins style="color: red; font-weight: bold; text-decoration: none;">1) We made S&Delta;TMD1 by deleting TMD1 of Sgene </ins></div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Reference===</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>===Reference===</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>----</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>----</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div># <html><a name="RefL001"></a></html>Rebecca White, Tram Anh T. Tran, Chelsey A. Dankenbring, John Deaton, and Ry Young.2010.The N-Terminal Transmembrane Domain of _ S Is Required for Holin but Not Antiholin Function_. J. Bacteriol. 192: 725–733 </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div># <html><a name="RefL001"></a></html>Rebecca White, Tram Anh T. Tran, Chelsey A. Dankenbring, John Deaton, and Ry Young.2010.The N-Terminal Transmembrane Domain of _ S Is Required for Holin but Not Antiholin Function_. J. Bacteriol. 192: 725–733 </div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div># <html><a name="RefL002"></a></html>CHUNG-YU CHANG, KIEBANG NAM, AND RY YOUNG1. 1995.2 S Gene Expression and the Timing of Lysis by Bacteriophage λ. J. Bacteriol. 177: 3283–3294</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div># <html><a name="RefL002"></a></html>CHUNG-YU CHANG, KIEBANG NAM, AND RY YOUNG1. 1995.2 S Gene Expression and the Timing of Lysis by Bacteriophage λ. J. Bacteriol. 177: 3283–3294</div></td></tr>
</table>Y.M.http://2010.igem.org/wiki/index.php?title=Team:Kyoto/Project/Goal_C&diff=204175&oldid=prevTomo: /* Construct */2010-10-28T01:29:34Z<p><span class="autocomment">Construct</span></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>It was used in the additional experiment, which is not finished.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>It was used in the additional experiment, which is not finished.</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>*<partinfo>BBa_K358019</partinfo>: Lysis cassette regulated by lacP, Lysis cassette[SRRz] is regulated by lactose promoter. </div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>*<partinfo>BBa_K358019</partinfo>: Lysis cassette regulated by lacP, Lysis cassette[SRRz] is regulated by lactose promoter. </div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>Activating lactose promoter and expressing SRRz gene, it causes the cell lysis. Thus, lactose promoter <del class="diffchange diffchange-inline">should </del>be repressed when <del class="diffchange diffchange-inline">transform </del>this part.</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>Activating lactose promoter and expressing SRRz gene, it causes the cell lysis. Thus, lactose promoter <ins class="diffchange diffchange-inline">must </ins>be repressed when this part <ins class="diffchange diffchange-inline">is transformed</ins>.</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>It was used in experiment 1 and 2.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>It was used in experiment 1 and 2.</div></td></tr>
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</table>Tomo