https://eln.excellgen.com/index.php?action=history&feed=atom&title=3257_pCMV5-anti-mkappa-rFc-His 3257 pCMV5-anti-mkappa-rFc-His - Revision history 2026-10-18T16:32:51Z Revision history for this page on the wiki MediaWiki 1.44.0 https://eln.excellgen.com/index.php?title=3257_pCMV5-anti-mkappa-rFc-His&diff=19453&oldid=prev Paul.xu at 21:29, 16 October 2018 2018-10-16T21:29:33Z <p></p> <p><b>New page</b></p><div>== &#039;&#039;&#039;Cloning of 3257 pCMV5-anti-mkappa-rFc-His&#039;&#039;&#039; ==<br /> <br /> &#039;&#039;&#039;Purpose:&#039;&#039;&#039; Cloning of 3257 pCMV5-anti-mkappa-rFc-His<br /> <br /> &#039;&#039;&#039;Date: &#039;&#039;&#039;08, 15, 2018<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: 3257&#039;&#039;&#039;<br /> <br /> &#039;&#039;&#039;Experimenters: &#039;&#039;&#039;yijia<br /> <br /> == &#039;&#039;&#039;Results&#039;&#039;&#039; ==<br /> <br /> === Cloning ===<br /> <br /> &#039;&#039;&#039;gene synthesis of 3257-F1&#039;&#039;&#039;<br /> <br /> &#039;&#039;&#039;template:&#039;&#039;&#039;<br /> &lt;div&gt;Gene synthesis&lt;/div&gt;&lt;div&gt;3257-1TGTCACGAATTCACAGGTGCAGCTGGTG&lt;/div&gt;&lt;div&gt;3257-2GTCTCAGAGAGCCGCCAGGTTGAACCCATCCTCCGCCAGATTCCACCAGCTGCACCTGTG&lt;/div&gt;&lt;div&gt;3257-3TGGCGGCTCTCTGAGACTGTCTTGTGCCGCCAGCGGCTTCACCTTTAGCGACACAGCCAT&lt;/div&gt;&lt;div&gt;3257-4CCCATTCTCTTCCCTTGCCAGGGGCCTGTCGGACCCACATCATGGCTGTGTCGCTAAAGG&lt;/div&gt;&lt;div&gt;3257-5GGCAAGGGAAGAGAATGGGTTGCCGCCATCGATACCGGCGGAGGCTACACCTACTACGCC&lt;/div&gt;&lt;div&gt;3257-6GCGTTGTCCCGGCTGATGGTGAATCTGCCCTTCACAGAATCGGCGTAGTAGGTGTAGCCT&lt;/div&gt;&lt;div&gt;3257-7TCAGCCGGGACAACGCCAAGAACACCCTGTACCTGCAGATGAACAGCCTGAAGCCTGAGG&lt;/div&gt;&lt;div&gt;3257-8AGTTGCCGCTGTAGGTCTTGGCGCAGTAGTACCGGGCTGTGTCCTCAGGCTTCAGGCTGT&lt;/div&gt;&lt;div&gt;3257-9AGACCTACAGCGGCAACTACTACAGCAACTACACCGTGGCCAACTACGGCACCACAGGCA&lt;/div&gt;&lt;div&gt;3257-10GGTACCGCTAGCGCTAGACACTGTGACCAGTGTTCCTCTGCCTGTGGTGCCGTAG<br /> [[File:IMG 1302.JPG|RTENOTITLE]]<br /> <br /> &#039;&#039;&#039;1st PCR of 3257-F2&#039;&#039;&#039;<br /> <br /> PCR&amp;nbsp; from&amp;nbsp; 1413 pCDNA3.4-Rabbit-IgG Heavy Chain&lt;br/&gt;3257-F2 CACTGGTCACAGTGTCTAGCGCTAGCGGTACCAGCAAGCCCACGTGCCCAC<br /> <br /> 3257-Not1-R3&amp;nbsp; TTGATTGCGGCCGCTCAGCAGTGATGGTGATGATGATGTTTACCCGGAGAGCGGGAGATG<br /> <br /> [[File:IMG 1311.JPG|RTENOTITLE]]<br /> <br /> &#039;&#039;&#039;2nd PCR of 3257&#039;&#039;&#039;<br /> <br /> combine both gene synthesis fragments and rabbit Fc fragment<br /> <br /> Purify with column PCR again using<br /> <br /> 3257-1 TGTCACGAATTCACAGGTGCAGCTGGTG<br /> <br /> 3257-Not1-R3 TTGATTGCGGCCGCTCAGCAGTGATGGTGATGATGATGTTTACCCGGAGAGCGGGAGATG<br /> <br /> 1160bp<br /> <br /> [[File:IMG 1312.JPG|RTENOTITLE]]<br /> <br /> &#039;&#039;&#039;Vector digestion&#039;&#039;&#039;<br /> <br /> Vector is 3075<br /> <br /> [[File:IMG 1304.JPG|RTENOTITLE]]<br /> &lt;/div&gt;<br /> === &amp;nbsp;Colony screen ===<br /> <br /> &#039;&#039;&#039;3257 PCR Screening&#039;&#039;&#039;<br /> <br /> [[File:IMG 1315.JPG|RTENOTITLE]]<br /> <br /> &#039;&#039;&#039;Miniprep of 3257&#039;&#039;&#039;<br /> <br /> {| border=&quot;1&quot; cellspacing=&quot;1&quot; cellpadding=&quot;1&quot; style=&quot;width:500px;&quot;<br /> |-<br /> | #<br /> | Lane<br /> | Enzymes<br /> | Length<br /> | Buffer<br /> | Note<br /> |-<br /> | 3257<br /> | 1-4(#1-4)<br /> | EcoR1+Not1<br /> | 1160bp<br /> | Fastdigest<br /> | #2 to do sequence<br /> |-<br /> | vector control<br /> | 3075<br /> | EcoR1+Not1<br /> | 681bp<br /> | Fastddigest<br /> | &lt;br/&gt;<br /> |}<br /> <br /> [[File:IMG 1316.JPG|RTENOTITLE]]<br /> <br /> 10/16/2018: in order to digest with Nhe1, plasmid has to be isolated from INV110.<br /> <br /> [[File:3257 3261 3260 INV110.JPG]]<br /> <br /> == &#039;&#039;&#039;Result intepretation/Summary&#039;&#039;&#039; ==<br /> <br /> == &#039;&#039;&#039;Conclusion&#039;&#039;&#039; ==<br /> <br /> &amp;lt;/div&amp;gt;<br /> [[Category:Cloning]]</div> Paul.xu