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Team:Bielefeld-CeBiTec/Notebook/Journal/Isobutanol/Sep
From 2014.igem.org
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<li><a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Protocols#RestrictionDigestion" target="_blank">Restriction digestion</a> with <a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/KitsAndEnzymes#RestrictionEnzymes" target="_blank"><i>EcoR</i>I</a> and <a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/KitsAndEnzymes#RestrictionEnzymes" target="_blank"><i>Pst</i>I</a> of pSB1A2_T7_<i>alsS</i>_<i>ilvC</i>_<i>ilvD</i>_<i>kivD</i> </li> | <li><a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Protocols#RestrictionDigestion" target="_blank">Restriction digestion</a> with <a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/KitsAndEnzymes#RestrictionEnzymes" target="_blank"><i>EcoR</i>I</a> and <a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/KitsAndEnzymes#RestrictionEnzymes" target="_blank"><i>Pst</i>I</a> of pSB1A2_T7_<i>alsS</i>_<i>ilvC</i>_<i>ilvD</i>_<i>kivD</i> </li> | ||
<ul> | <ul> | ||
| - | <li>Bands not as expected (~ | + | <li>Bands not as expected (~7000bp)</li> |
</ul> </ul></ul></ul></ul> | </ul> </ul></ul></ul></ul> | ||
| + | <ul> | ||
| + | <li><b><i>pSB1K3_alsS_ilvC_ilcD_kivD_adhA</i></b></li> | ||
| + | <ul> | ||
| + | <li>This week we tried to combine the pSB1K3_<i>alsS</i>_<i>ilvC</i>_<i>ilvD</i>_<i>kivD</i> with <i>adhA</i></li> | ||
| + | <ul> | ||
| + | <li><a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Protocols#BioBrick" target="_blank">BioBrick Assembly</a> (Suffix)</li> | ||
| + | <ul> | ||
| + | <li>Backbone (digested with <a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/KitsAndEnzymes#RestrictionDigestion" target="_blank"><i>Spe</i>I</a>, <a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/KitsAndEnzymes#RestrictionDigestion" target="_blank"><i>Pst</i>I</a>)</li> | ||
| + | <ul> | ||
| + | <li>pSB1K3_<i>alsS</i>_<i>ilvC</i>_<i>ilvD</i>_<i>kivD</i></li> | ||
| + | </ul> | ||
| + | <li>Insert (digested with <a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/KitsAndEnzymes#RestrictionDigestion" target="_blank"><i>Xba</i>I</a>, <a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/KitsAndEnzymes#RestrictionDigestion" target="_blank"><i>Pst</i>I</a>)</li> | ||
| + | <ul> | ||
| + | <li><i>pSB1K3_<i>alsS</i>_<i>ilvC</i>_<i>ilvD</i>_<i>kivD</i>_<i>adhA</i></i></li> | ||
| + | </ul> | ||
| + | </ul> | ||
| + | </ul><ul> | ||
| + | <li><a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Protocols#ColonyPCR" target="_blank">Colony PCR</a> (<a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Primer#fw-ilvD-kivD" target="_blank">fw-ilvD-kivD</a>, <a href="https://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Primer#rev-pSB1C3-adhA" target="_blank">rev-pSB1C3-adhA</a>) | ||
| + | </li> | ||
| + | <ul> | ||
| + | <li>Annealing temperature: 65 °C</li> | ||
| + | <li>Bands not as expected (~ 3800 bp)</li> | ||
| + | </ul> | ||
| + | </ul> | ||
| + | |||
| + | |||
| + | </ul> | ||
| + | </ul> | ||
| + | <br> | ||
| + | |||
</div> | </div> | ||
</div> | </div> | ||
Revision as of 10:27, 15 October 2014
 |
September |
 |
- adhA
- This week we tried to verify the positive cutures that were identified last week by a restriction digest
- Plasmid isolation of pSB1K3_adhA
- Restriction digestion with EcoRI and PstI of pSB1K3_adhA
- Bands as expected (~1200 bp)
- Successful sequencing
Agarose gel from the restriction digestion with EcoRI and PstI. As a Ladder we used GeneRuler™ 1 kb DNA Ladder from Thermo Scientific.
- T7_alsS_ilvC_ilvD_kivD
- This week we tried to verify the positive cutures that were identified last week by a restriction digest
- Plasmid isolation of pSB1A2_T7_alsS_ilvC_ilvD_kivD
- Restriction digestion with EcoRI and PstI of pSB1A2_T7_alsS_ilvC_ilvD_kivD
- Bands not as expected (~7000bp)
- pSB1K3_alsS_ilvC_ilcD_kivD_adhA
- This week we tried to combine the pSB1K3_alsS_ilvC_ilvD_kivD with adhA
- BioBrick Assembly (Suffix)
- Colony PCR (fw-ilvD-kivD, rev-pSB1C3-adhA)
- Annealing temperature: 65 °C
- Bands not as expected (~ 3800 bp)
