Difference between revisions of "Team:UMaryland/Notebook"

Line 287: Line 287:
 
Miraculin
 
Miraculin
 
<ul class="a">
 
<ul class="a">
−
   <li>- RE digest on β-cyclase, AppY, CREB, and CRTBEY genes along with the pSB1C3 backbone and the pBAD + miraculin out of the PSB1A3 backbone so that we could move the genes to the pSB1C3 backbone</li>
+
   <li>- RE digest on β-cyclase, AppY, CREB, and CRTBEY genes along with the pSB1C3 backbone and the pBAD + miraculin out of the pSB1A3 backbone so that we could move the genes to the pSB1C3 backbone</li>
 
   <li>- Ran gel with all parts and cut out the bands</li>
 
   <li>- Ran gel with all parts and cut out the bands</li>
 
   <li>- Mini-prepped SRNBC + constitutive colicinFy and shipped them off to be sequenced</li>
 
   <li>- Mini-prepped SRNBC + constitutive colicinFy and shipped them off to be sequenced</li>
 
   <li>- Performed gel purification on gels from day before but they failed; they didn’t show up in the spectrophotometer</li>
 
   <li>- Performed gel purification on gels from day before but they failed; they didn’t show up in the spectrophotometer</li>
−
   <li>- Performed minipreps on pBAD+Miraculin in the PSB1C3 backbone as well as the const. GFP+SRNBC</li>
+
   <li>- Performed minipreps on pBAD+Miraculin in the pSB1C3 backbone as well as the const. GFP+SRNBC</li>
 
   <li>- Performed RE digests on the previous mini-preps: EcoRI and PstI on pBAD + Miraculin, and XBa1 and Pst1 on SRNBC + Constitutive GFP</li>
 
   <li>- Performed RE digests on the previous mini-preps: EcoRI and PstI on pBAD + Miraculin, and XBa1 and Pst1 on SRNBC + Constitutive GFP</li>
 
</ul>
 
</ul>
Line 326: Line 326:
 
   <li>- Performed transformations of the constitutive GFP + SRNBC construct</li>
 
   <li>- Performed transformations of the constitutive GFP + SRNBC construct</li>
 
   <li>- Transformations of SRNBC + Constitutive GFP failed</li>
 
   <li>- Transformations of SRNBC + Constitutive GFP failed</li>
−
   <li>- Performed PCR on the PSB1C3 backbone in order to amplify it in preparation for a Gibson Assembly with the Hok/Sok gene</li>
+
   <li>- Performed PCR on the pSB1C3 backbone in order to amplify it in preparation for a Gibson Assembly with the Hok/Sok gene</li>
 
</ul>
 
</ul>
 
<br>
 
<br>
Line 358: Line 358:
 
Hok/Sok
 
Hok/Sok
 
<ul class="a">
 
<ul class="a">
−
   <li>- Performed an RE digest on the PSB1C3 backbone with XbaI and SpeI and then ran it through a gel to separate it out, to prepare for the moving of Hok/Sok into pSB1C3</li>
+
   <li>- Performed an RE digest on the pSB1C3 backbone with XbaI and SpeI and then ran it through a gel to separate it out, to prepare for the moving of Hok/Sok into pSB1C3</li>
 
</ul>
 
</ul>
 
<br>
 
<br>
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Hok/Sok
 
Hok/Sok
 
<ul class="a">
 
<ul class="a">
−
   <li>- inserted Hok/Sok gblock into PSB1C3 backbone using Gibson assembly</li>
+
   <li>- inserted Hok/Sok gblock into pSB1C3 backbone using Gibson assembly</li>
 
   <li>- construct sent for sequencing </li>
 
   <li>- construct sent for sequencing </li>
 
</ul>
 
</ul>
Line 424: Line 424:
 
Interlab
 
Interlab
 
<ul class="a">
 
<ul class="a">
−
   <li>- performed a 3A assembly of each promoter + GFP in PSB1K3 (GFP in PSB1A3 and promoters in PSB1C3) </li>
+
   <li>- performed a 3A assembly of each promoter + GFP in PSB1K3 (GFP in pSB1A3 and promoters in pSB1C3) </li>
 
</ul>
 
</ul>
 
<br>
 
<br>
Line 477: Line 477:
 
   <li>- const_promoter + RBS + RFP were replated from last week but produced no colonies </li>
 
   <li>- const_promoter + RBS + RFP were replated from last week but produced no colonies </li>
 
   <li>- re-transformed original and new 3A assembly which produced the correct sequence  </li>
 
   <li>- re-transformed original and new 3A assembly which produced the correct sequence  </li>
−
   <li>- ligated const_promoter:QD-RFP in PSB1C3 </li>
+
   <li>- ligated const_promoter:QD-RFP in pSB1C3 </li>
−
   <li>- performed a 3A assembly of Hok/Sok plasmid with QD-RFP downstream in PSB1K3 </li>
+
   <li>- performed a 3A assembly of Hok/Sok plasmid with QD-RFP downstream in pSB1K3 </li>
 
   <li>- site directed mutagenesis of quick degrading GFP (Bba_K750000)  failed </li>
 
   <li>- site directed mutagenesis of quick degrading GFP (Bba_K750000)  failed </li>
 
<ul class="a">
 
<ul class="a">
Line 587: Line 587:
 
<ul class="a">
 
<ul class="a">
 
   <li>- 3A assembly of H/S + RFP failed </li>
 
   <li>- 3A assembly of H/S + RFP failed </li>
−
   <li>- ran a PCR of PSB1C3+H/S+RFP using 3 different rxn buffers  </li>
+
   <li>- ran a PCR of pSB1C3+H/S+RFP using 3 different rxn buffers  </li>
 
   <li>- HF rxn buffer </li>
 
   <li>- HF rxn buffer </li>
 
   <li>- GC rxn buffer </li>
 
   <li>- GC rxn buffer </li>
Line 600: Line 600:
 
   <li>- ran PCR for GFP 1, GFP 3, and IL1 which produced the correct sized products  </li>
 
   <li>- ran PCR for GFP 1, GFP 3, and IL1 which produced the correct sized products  </li>
 
   <li>- Gibson Assembly of GFP + IL1 produced colonies </li>
 
   <li>- Gibson Assembly of GFP + IL1 produced colonies </li>
−
   <li>- PSB1C3 - IL3 did not have bands in gel of appropriate size  </li>
+
   <li>- pSB1C3 - IL3 did not have bands in gel of appropriate size  </li>
  
 
</ul>
 
</ul>
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Lutein
 
Lutein
 
<ul class="a">
 
<ul class="a">
−
   <li>- RE digests of pLAC-RFP in PSB1C3 </li>
+
   <li>- RE digests of pLAC-RFP in pSB1C3 </li>
 
   <li>- ran a Gibson Assembly of E-cyclase/C3 and E-hydroxylase/C3 which produced colonies  </li>
 
   <li>- ran a Gibson Assembly of E-cyclase/C3 and E-hydroxylase/C3 which produced colonies  </li>
 
</ul>
 
</ul>

Revision as of 03:51, 19 September 2015