Team:Goettingen/week10-2

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#2 Speed Improvement - 10th week

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V07_02

V07_02_1 Preparative double digestion of 20E-flhDC, 2G-flhDC, 18O-flhDC and pSB1C3

Experiment:
In order to clone the flhDC-promoter constructs intro pSB1C3, all components were digested with EcoRI and PstI according to the protocol. The restrictio products were subsequently separated on a gel in order to verify the digestions success and purification of the desired fragments.

Observations & Results:
The gel did not feature fragments of the expected size.Obviously, something was wrong with the digestion.

V07_02_2 Preparation of over night cultures

Experiment:
In order to gain more plasmid material of the flhDC-promoter constructs over night cultured were prepared for a subsequent plasmid isolation.
20E - #2
20G - #1
2G - #1
20I - #2
18M - #2
18O - #2
18K - #1
18C - #2

V07_03

V07_03_1 Miniprep of the flhDC-promoter constructs

Experiment:
Minipreps were conducted with PeqGOLD MiniPrep Kit (Peqlab) according to the user manual.

V07_03_2 Amplification of the genes fliC (DH10B), fliC (Salmonella), motA, motB and yhjH

Experiment:
The genes fliC (DH10B), fliC (Salmonella), motA, motB and yhjH were amplified by Pfu-Polymerase according to the following protocol. The success of the PCR was subsequently investigated preparing a 1% agarose gel.
Observations & Results:
The PCR failed for no bands were visible on the gels whereas the marker was clearly discernible.

V06_14

Preparation of over night cultures

Experiment:
Over night cultures of the pBAD-sfGFP clones were prepared in order to isolate the plasmids.

V06_15

Miniprep of pBAD-sfGFP

Experiment:
Minipreps were conducted with PeqGOLD MiniPrep Kit (Peqlab) according to the user manual.

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@@ Line 550: / Line 550: @@
 C - #2 <br></li>
      </ul>
-<br></td></tr>
-</table>
-<br>
-<table cellpadding="20 px" border="1" bordercolor="black" valign="top">
-<tr bordercolor="black" valign="top">
-<td width="900" bordercolor="black" valign="top">
-<h2><b>V06_12 </b></h2><br>
-<b>Preparation of over night cultures</i></b><br>
-<ul>
-<li>Experiment: <br>
-In order to gain further plasmid material over night cultures of one colonie of all eight <i>flhDC</i>-promoter constructs were prepared for subsequent plasmid isolation. <br>
-E - #2 <br>
-G - #1 <br>
-G - #1 <br>
-I - #2 <br>
-M - #2 <br>
-O - #2 <br>
-K - #1 <br>
-C - #2 <br>
 <br></td></tr>
 </table>
@@ Line 580: / Line 559: @@
 <td width="900" bordercolor="black" valign="top">
-<h2><b>V06_13 </b></h2><br>
+<h2><b>V07_03 </b></h2><br>
-<b>V06_13_1 Miniprep of the new <i>flhDC</i>-promoter constructs</b><br>
+<b>V07_03_1 Miniprep of the <i>flhDC</i>-promoter constructs</b><br>
 <ul>
 <li>Experiment: <br>
 Minipreps were conducted with PeqGOLD MiniPrep Kit (Peqlab) according to the user manual.</li></ul>
 <br>
-<b>V06_13_2 Chemical transformartion of pBAD-<i>sfGFP</i> into <i>E. coli</i> (DH10B)</b><br>
+<b>V07_03_2 Amplification of the genes <i>fliC</i> (DH10B), <i>fliC</i> (Salmonella), <i>motA</i>, <i>motB</i> and <i>yhjH</i></b><br>
 <ul>
 <li>Experiment:  <br>
-For the chemical transformation the standard protocol was followed. <br>
+The genes <i>fliC</i> (DH10B), <i>fliC</i> (Salmonella), <i>motA</i>, <i>motB</i> and <i>yhjH</i> were amplified by Pfu-Polymerase according to the following protocol. The success of the PCR was subsequently investigated preparing a 1% agarose gel.<br>
 <li>Observations & Results: <br>
-The transformation was successful since all plates showed numeours colonoes except the negative control.</li>
+The PCR failed for no bands were visible on the gels whereas the marker was clearly discernible.</li>
 </ul><br>