https://eln.excellgen.com/index.php?action=history&feed=atom&title=1002-pLysN 2026-10-18T08:04:12Z Revision history for this page on the wiki MediaWiki 1.44.0 https://eln.excellgen.com/index.php?title=1002-pLysN&diff=13956&oldid=prev 2015-02-25T16:09:52Z

<p></p> <p><b>New page</b></p><div>== &#039;&#039;&#039;Cloning of 1002-pLysN&#039;&#039;&#039; ==<br /> <br /> &#039;&#039;&#039;Purpose:&#039;&#039;&#039; Cloning of 1002-pLysN by PCR <br /> <br /> &#039;&#039;&#039;Date: &#039;&#039;&#039;2012-2-10--2012-2-17 <br /> <br /> &#039;&#039;&#039;Notebook:003737&#039;&#039;&#039; <br /> <br /> &#039;&#039;&#039;Page:&#039;&#039;&#039; <br /> <br /> &#039;&#039;&#039;Clone ID:1002&#039;&#039;&#039; <br /> <br /> &#039;&#039;&#039;Experimenters:&#039;&#039;&#039; yingying <br /> <br /> == &#039;&#039;&#039;Results&#039;&#039;&#039; ==<br /> <br /> === cloning ===<br /> <br /> Date:2012-2-10 <br /> <br /> [[Image:1002 PCR 2-14-2012.JPG]]&lt;br&gt; <br /> <br /> 1. PCR LacZProm-Sal1-F + LacZProm-R using pUC19 as template. 15 cycles&lt;br&gt;2. PCR T7LysN-F + T7LysN-BamH1-R, using pLysS as template, 15 cycles&lt;br&gt;pLysS plasmid can be extracted from BL21(DE3) pLysS&lt;br&gt;3. DO not run gel, Mix two PCR product, Purify PCR product using column, elute with 20 ul water.&lt;br&gt;4. PCR using LacZProm-Sal1-F + T7LysN-BamH1-R using step 3 eluate as template (all ~ 20 ul),25 cycles&lt;br&gt; <br /> <br /> [[Image:546+1002+1005 clone 2-15-2012.JPG]]&lt;br&gt; <br /> <br /> {| cellspacing=&quot;1&quot; cellpadding=&quot;1&quot; border=&quot;1&quot; style=&quot;width: 334px; height: 64px;&quot;<br /> |-<br /> | &lt;br&gt; <br /> | band 1&lt;br&gt; <br /> | band 2&lt;br&gt;<br /> |-<br /> | &lt;br&gt; <br /> | PCR product&lt;br&gt; <br /> | pACYC184 1003&lt;br&gt;<br /> |-<br /> | digest enzymes&lt;br&gt; <br /> | colspan=&quot;2&quot; | &amp;nbsp;&amp;nbsp;&amp;nbsp;&amp;nbsp; Sal1 + Bamh1&lt;br&gt;<br /> |}<br /> <br /> &lt;br&gt; <br /> <br /> {| cellspacing=&quot;1&quot; cellpadding=&quot;1&quot; border=&quot;1&quot;<br /> |-<br /> | &lt;br&gt;<br /> |}<br /> <br /> === Colony Screening ===<br /> <br /> Date: 2012-2-17 <br /> <br /> Make miniprep and digest with&amp;nbsp; Sal1 + Bamh1 <br /> <br /> [[Image:1002 digest detect 2-17-2012.JPG]] <br /> <br /> clone 1,2,3,5,6 may be right, need to sequence <br /> <br /> &lt;br&gt; <br /> <br /> &lt;br&gt; <br /> <br /> &lt;br&gt; &lt;br&gt; <br /> <br /> == &#039;&#039;&#039;Result intepretation/Summary&#039;&#039;&#039; ==<br /> <br /> *&amp;lt;br&amp;gt;<br /> <br /> == &#039;&#039;&#039;Conclusion&#039;&#039;&#039; ==<br /> <br /> &amp;lt;br&amp;gt; <br /> <br /> {| cellspacing=&quot;1&quot; cellpadding=&quot;1&quot;<br /> |-<br /> | &lt;br&gt;<br /> |}<br /> <br /> &amp;lt;br&amp;gt; <br /> <br /> {| cellspacing=&quot;1&quot; cellpadding=&quot;1&quot;<br /> |-<br /> | &lt;br&gt;<br /> |}<br /> <br /> &lt;br&gt; <br /> <br /> {| cellspacing=&quot;1&quot;<br /> |-<br /> | &lt;br&gt;<br /> |}<br /> <br /> == &#039;&#039;&#039;Cloning of 1002-pLysN&#039;&#039;&#039; ==<br /> <br /> &#039;&#039;&#039;Purpose:&#039;&#039;&#039; Cloning of 1002-pLysN by PCR <br /> <br /> &#039;&#039;&#039;Date: &#039;&#039;&#039;2012-2-22--2012-2-27 <br /> <br /> &#039;&#039;&#039;Notebook:003737&#039;&#039;&#039; <br /> <br /> &#039;&#039;&#039;Page:&#039;&#039;&#039; <br /> <br /> &#039;&#039;&#039;Clone ID:1002&#039;&#039;&#039; <br /> <br /> &#039;&#039;&#039;Experimenters:&#039;&#039;&#039; yingying <br /> <br /> == &#039;&#039;&#039;Results&#039;&#039;&#039; ==<br /> <br /> === cloning ===<br /> <br /> Date:2012-2-10 <br /> <br /> [[Image:1002 PCR 2-14-2012.JPG]]&lt;br&gt; <br /> <br /> 1. PCR LacZProm-Sal1-F + LacZProm-R using pUC19 as template. 15 cycles&lt;br&gt;2. PCR T7LysN-F + T7LysN-BamH1-R, using pLysS as template, 15 cycles&lt;br&gt;pLysS plasmid can be extracted from BL21(DE3) pLysS&lt;br&gt;3. DO not run gel, Mix two PCR product, Purify PCR product using column, elute with 20 ul water.&lt;br&gt;4. PCR using LacZProm-Sal1-F + T7LysN-BamH1-R using step 3 eluate as template (all ~ 20 ul),25 cycles&lt;br&gt; <br /> <br /> [[Image:1002_clone_2-22-2012.JPG]]&lt;br&gt; <br /> <br /> {| cellspacing=&quot;1&quot; cellpadding=&quot;1&quot; border=&quot;1&quot; style=&quot;width: 334px; height: 64px;&quot;<br /> |-<br /> | &lt;br&gt; <br /> | band 1&lt;br&gt; <br /> | band 2&lt;br&gt;<br /> |-<br /> | &lt;br&gt; <br /> | PCR product&lt;br&gt; <br /> | pACYC184 1003&lt;br&gt;<br /> |-<br /> | digest enzymes&lt;br&gt; <br /> | colspan=&quot;2&quot; | &amp;nbsp;&amp;nbsp;&amp;nbsp;&amp;nbsp; Sal1 + Bamh1&lt;br&gt;<br /> |}<br /> <br /> &lt;br&gt; <br /> <br /> {| cellspacing=&quot;1&quot; cellpadding=&quot;1&quot; border=&quot;1&quot;<br /> |-<br /> | &lt;br&gt;<br /> |}<br /> <br /> === Colony Screening ===<br /> <br /> Date: 2012-2-24 <br /> <br /> Make miniprep and digest with&amp;nbsp; Sal1 + Bamh1 <br /> <br /> [[Image:1002_digest_detect_2-24-2012.JPG]]&lt;br&gt; <br /> <br /> clone 1,2,3,5,6 may be right, need to sequence <br /> <br /> &lt;br&gt; <br /> <br /> &lt;br&gt; <br /> <br /> &lt;br&gt; &lt;br&gt; <br /> <br /> == &#039;&#039;&#039;Result intepretation/Summary&#039;&#039;&#039; ==<br /> <br /> *&amp;lt;br&amp;gt;<br /> <br /> == &#039;&#039;&#039;Conclusion&#039;&#039;&#039; ==<br /> <br /> <br /> <br /> [[Category:Cloning]]</div>

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