https://eln.excellgen.com/index.php?action=history&feed=atom&title=1250_pET28b-KOD-2 1250 pET28b-KOD-2 - Revision history 2026-10-18T07:29:31Z Revision history for this page on the wiki MediaWiki 1.44.0 https://eln.excellgen.com/index.php?title=1250_pET28b-KOD-2&diff=5042&oldid=prev Paul.xu at 19:47, 10 July 2012 2012-07-10T19:47:42Z <p></p> <p><b>New page</b></p><div>== &#039;&#039;&#039;Cloning of 1250 pET28b-KOD-2&#039;&#039;&#039; ==<br /> <br /> &#039;&#039;&#039;Purpose:&#039;&#039;&#039; Cloning of 1250 pET28b-KOD-2 <br /> <br /> &#039;&#039;&#039;Date: &#039;&#039;&#039;July 6, 2012&lt;br&gt; <br /> <br /> &#039;&#039;&#039;Notebook:&amp;nbsp;&#039;&#039;&#039; <br /> <br /> &#039;&#039;&#039;Page:&#039;&#039;&#039; <br /> <br /> &#039;&#039;&#039;Clone ID: &#039;&#039;&#039;1250 <br /> <br /> &#039;&#039;&#039;Experimenters: &#039;&#039;&#039;Jennifer Truong <br /> <br /> == &#039;&#039;&#039;Results&#039;&#039;&#039; ==<br /> <br /> === Cloning ===<br /> <br /> Date:&amp;nbsp;July 7, 2012&lt;br&gt; <br /> <br /> Digest 480 pET28b-KOD with EcoR1 + Not1, see lane 5 <br /> <br /> Cut out and purify fragment <br /> <br /> [[Image:0706 digestion.JPG]]&lt;br&gt;PCR this ~120bp fragment out from 480 pET28b-KOD using primer&lt;br&gt;KOD-EcoR1-F TTGAGA GAATTC TGAGAGCCTTC&lt;br&gt;KOD-Not1-R GGATCCGCGGCCGCGGTACCAGTTCCCTTCGGCTTCAGCCAAG&lt;br&gt;[[Image:1250 amp.JPG]]&lt;br&gt; <br /> <br /> Expected size ~120 bp, cut out band, elude with 50 uL&amp;nbsp;water&lt;br&gt; <br /> <br /> Digest with EcoR1 + Not1&lt;br&gt;clone back into 480 pET28b-KOD &lt;br&gt; <br /> <br /> [[Image:0707 digestion.JPG]]&lt;br&gt; <br /> <br /> Digested pET28b-KOD (lane 6)&amp;nbsp;and digested PCRed pET28b-KOD (lane 5), saved vector from lane 6, but need to rePCR the digested pET28b-KOD&lt;br&gt; Date:&amp;nbsp;June 9, 2012&lt;br&gt; <br /> <br /> [[Image:0709 pcr.JPG]] <br /> <br /> Expected size ~120 bp (see lane 4), cut out band, purify&lt;br&gt; <br /> <br /> Digest with EcoR1 + Not1&lt;br&gt;clone back into 480 pET28b-KOD &lt;br&gt; <br /> <br /> [[Image:1250_digestion.JPG]]&lt;br&gt;<br /> <br /> Digest DNA (lane 1) and vector 480 (lane 2) with EcoR1 + Not1&lt;br&gt;Clone back into 480 pET28b-KOD &lt;br&gt;<br /> <br /> Cut out bands in lanes 1 and 2, purify together and ligate<br /> <br /> Transform with Top10 and plate on Kan plate, grow overnight<br /> <br /> === Colony Screening&lt;br&gt; ===<br /> <br /> Date:&amp;nbsp;July 10, 2012<br /> <br /> [[Image:1250_Screening.JPG]]<br /> <br /> Only samples 5 and 6 had positive inserts<br /> <br /> == &#039;&#039;&#039;Result intepretation/Summary&#039;&#039;&#039;&lt;br&gt; ==<br /> <br /> == &#039;&#039;&#039;Conclusion&#039;&#039;&#039;&lt;br&gt; ==<br /> <br /> [[Category:Cloning]]</div> Paul.xu