Team:SYSU-China/project/cyclecontrol
From 2012.igem.org
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- | <td width="286" height=" | + | <td width="286" height="152"> </td> |
- | <td width="76" align="center" valign="middle"> | + | <td width="76" align="center" valign="middle"><p class="style1"><a href="https://2012.igem.org/Team:SYSU-China/project">Background</a></p></td> |
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<td width="20"> </td> | <td width="20"> </td> | ||
- | <td width="72" align="center" valign="middle"> | + | <td width="72" align="center" valign="middle"><p class="style3">Cycle Control</p></td> |
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<td width="15"> </td> | <td width="15"> </td> | ||
- | <td width="73" align="center" valign="middle"> | + | <td width="73" align="center" valign="middle"><p><a href="https://2012.igem.org/Team:SYSU-China/project/quorumtransform" class="style1">Quorum Transform</a><a href="https://2012.igem.org/Team:SYSU-China/project/industrialapplication" class="style1"></a></p></td> |
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<td width="14"> </td> | <td width="14"> </td> | ||
- | <td width="76" align="center" valign="middle"> | + | <td width="76" align="center" valign="middle"><p><a href="https://2012.igem.org/Team:SYSU-China/project/theoreticalvalue" class="style1">Theoretical Valuel</a></p></td> |
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<td width="12"> </td> | <td width="12"> </td> | ||
- | <td width="84" align="center" valign="middle"> | + | <td width="84" align="center" valign="middle"><p><a href="https://2012.igem.org/Team:SYSU-China/project/manufactureapplications" class="style1">Manufacture Applications</a></p></td> |
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<td width="14"> </td> | <td width="14"> </td> | ||
- | <td width="72" align="center" valign="middle"> | + | <td width="72" align="center" valign="middle"><p><a href="https://2012.igem.org/Team:SYSU-China/project/supergeneguard" class="style1">Super Geneguard</a></p></td> |
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</tr> | </tr> | ||
</table> | </table> | ||
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<td width="46" rowspan="8"> </td> | <td width="46" rowspan="8"> </td> | ||
<td width="612" rowspan="8" align="center" valign="middle"><div style="height:590px;width:612px;overflow:scroll"> | <td width="612" rowspan="8" align="center" valign="middle"><div style="height:590px;width:612px;overflow:scroll"> | ||
- | <h3 | + | <h3 align="left">Introduction</h3> |
- | + | <p align="left">We can regulate bacteria cell cycle by controlling DNA replication, RNA transcription, or protein translation, but the regulation of DNA replication initiation is promptest and do the less harm ro basic metablism.</p> | |
- | <p>We can regulate bacteria cell cycle by controlling DNA replication, RNA transcription, or protein translation, but the regulation of DNA replication initiation is promptest and do the less harm ro basic metablism.</p> | + | <p align="left">DNA replication in bacteria is controlled at the level of initiation. Bidirectional chromosomal replication from the unique replication origin, oriC, towards the terminus, terC, requires a constant time at a given temperature and assisted by lots of proteins. [1]</p> |
- | <p>DNA replication in bacteria is controlled at the level of initiation. Bidirectional chromosomal replication from the unique replication origin, oriC, towards the terminus, terC, requires a constant time at a given temperature and assisted by lots of proteins. [1]</p> | + | <h3 align="left">Method</h3> |
- | <h3>Method</h3> | + | <p align="left"><u>Plasmid Construction</u></p> |
- | <p><u>Plasmid Construction</u></p> | + | <p align="left">Gene iciA, hda, seqA, cspD were cloned from E.coli strain BL21, and link to RFP. We test the function of the four genes in vector pUC18 respectively.</p> |
- | <p>Gene iciA, hda, seqA, cspD were cloned from E.coli strain BL21, and link to RFP. We test the function of the four genes in vector pUC18 respectively.</p> | + | |
<p align="center"><img src="https://static.igem.org/mediawiki/2012/3/3a/SYSU-plasmidconstruction.jpg" alt="" width="500" height="618" /></p> | <p align="center"><img src="https://static.igem.org/mediawiki/2012/3/3a/SYSU-plasmidconstruction.jpg" alt="" width="500" height="618" /></p> | ||
<p align="center">img.1 gel electrophoresis of 4 cycle regulative genes (A: PCR 4 genes from genomic DNA of BL21strain with self-designed primers; B: connection 4 gened with RFP respectively then link to pUC18 vectors.)</p> | <p align="center">img.1 gel electrophoresis of 4 cycle regulative genes (A: PCR 4 genes from genomic DNA of BL21strain with self-designed primers; B: connection 4 gened with RFP respectively then link to pUC18 vectors.)</p> | ||
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<p align="left"><u>Growth</u></p> | <p align="left"><u>Growth</u></p> | ||
<ol> | <ol> | ||
- | <li class="style4">Transform the vectors into E.coli strain DH5α as to amplify vectors and measure the specific parameters of cycle delay. Shake the bacteria in 50mL LB liquid in 37℃, 260rpm. Extract 100ul Bacteria Solution and store on 96-well plate in 4℃ every two hours for protein accumulation test. Extract 100ul Bacteria Solution and measure bacteria concentration through OD600 every hour. *IPTG was added in OD600=0.2, final concentration=0.5mL</li> | + | <li class="style4"> |
- | <li class="style4">2) Transform the vectors into E.coli strain GE in whose chromosome GFP gene was integrated. Shake the bacteria in 50mL lactose liquid media in 37℃, 260rpm. Extract triplicate of 100ul Bacteria Solution and store on 96-well plate in 4℃ every two hours for measuring protein production, energy consumption, and metabolic efficiency. *IPTG was added in OD600=0.2, final concentration=0.5mL</li> | + | <div align="left">Transform the vectors into E.coli strain DH5α as to amplify vectors and measure the specific parameters of cycle delay. Shake the bacteria in 50mL LB liquid in 37℃, 260rpm. Extract 100ul Bacteria Solution and store on 96-well plate in 4℃ every two hours for protein accumulation test. Extract 100ul Bacteria Solution and measure bacteria concentration through OD600 every hour. *IPTG was added in OD600=0.2, final concentration=0.5mL</div> |
+ | </li> | ||
+ | <li class="style4"> | ||
+ | <div align="left">2) Transform the vectors into E.coli strain GE in whose chromosome GFP gene was integrated. Shake the bacteria in 50mL lactose liquid media in 37℃, 260rpm. Extract triplicate of 100ul Bacteria Solution and store on 96-well plate in 4℃ every two hours for measuring protein production, energy consumption, and metabolic efficiency. *IPTG was added in OD600=0.2, final concentration=0.5mL</div> | ||
+ | </li> | ||
</ol> | </ol> | ||
- | <h3>Result</h3> | + | <h3 align="left">Result</h3> |
- | <p><u>Bacteria Cell Growth Rate</u></p> | + | <p align="left"><u>Bacteria Cell Growth Rate</u></p> |
<p align="center"><img src="https://static.igem.org/mediawiki/2012/4/45/SYSU-Growthrate.png" alt="" width="583" height="349" /></p> | <p align="center"><img src="https://static.igem.org/mediawiki/2012/4/45/SYSU-Growthrate.png" alt="" width="583" height="349" /></p> | ||
<p align="center">img.2 Curve of bacteria concentration in OD600 by hours(time range from 0 to 12 hour. A:iciA, B:hda, C:seqA, D:cspD)</p> | <p align="center">img.2 Curve of bacteria concentration in OD600 by hours(time range from 0 to 12 hour. A:iciA, B:hda, C:seqA, D:cspD)</p> |
Revision as of 08:53, 26 September 2012