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Team:TU Darmstadt/Notebook/Labjournal/K1497007
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| - | <!--TYPO3SEARCH_begin--><div id="c116" class="csc-default"><div class="csc-header csc-header-n1"><h1 class="csc-firstHeader">K1497007 - Naringenin operon</h1></div><div class="csc-textpic-text"><div><p>For combing all for four genes of our operon, we used the previously obtained Biobricks B0034-CHS-B0034-CHI-pSB1C3 (K1497005) and B0034-4CL-B0034-TAL-pSB1C3 (K1497006). We digested B0034-4CL-B0034-TAL-pSB1C3 with EcoRI + SpeI and B0034-CHS-B0034-CHI-pSB1C3 with EcoRI + XbaI. Furthermore we dephosphorylated the digestion of B0034-CHS-B0034-CHI-pSB1C3. We ligated both digestions with T4 ligase and transformed the products into Top10 E.coli via heat shock. We spread out on CMP-LB agar plates and incubated the plates overnight at 37°C. We analyzed obtained colonies by colony PCR and agarose gel electrophoresis. Positive colonies were inoculated in 5 ml LB with CMP and plasmids were isolated after incubating for 16 h at 37°C. For further verification we send the samples to Eurofins for sequencing. </p></div></div></div><!--TYPO3SEARCH_end--> | + | <!--TYPO3SEARCH_begin--><div id="c116" class="csc-default"><div class="csc-header csc-header-n1"><h1 class="csc-firstHeader">K1497007 - Naringenin operon</h1></div><div class="csc-textpic-text"><div><p>For combing all for four genes of our operon, we used the previously obtained Biobricks B0034-CHS-B0034-CHI-pSB1C3 (K1497005) and B0034-4CL-B0034-TAL-pSB1C3 (K1497006). We digested B0034-4CL-B0034-TAL-pSB1C3 with EcoRI + SpeI and B0034-CHS-B0034-CHI-pSB1C3 with EcoRI + XbaI. Furthermore we dephosphorylated the digestion of B0034-CHS-B0034-CHI-pSB1C3. We ligated both digestions with T4 ligase and transformed the products into Top10 E.coli via heat shock. We spread out on CMP-LB agar plates and incubated the plates overnight at 37°C. We analyzed obtained colonies by colony PCR and agarose gel electrophoresis. Positive colonies were inoculated in 5 ml LB with CMP and plasmids were isolated after incubating for 16 h at 37°C gaining the BioBrick <a href="http://parts.igem.org/Part:BBa_K1497007" title="Opens internal link in current window" class="internal-link"> BBa_K1497007</a>. For further verification we send the samples to Eurofins for sequencing. </p></div></div></div><!--TYPO3SEARCH_end--> |
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Latest revision as of 18:50, 17 October 2014
K1497007 - Naringenin operon
For combing all for four genes of our operon, we used the previously obtained Biobricks B0034-CHS-B0034-CHI-pSB1C3 (K1497005) and B0034-4CL-B0034-TAL-pSB1C3 (K1497006). We digested B0034-4CL-B0034-TAL-pSB1C3 with EcoRI + SpeI and B0034-CHS-B0034-CHI-pSB1C3 with EcoRI + XbaI. Furthermore we dephosphorylated the digestion of B0034-CHS-B0034-CHI-pSB1C3. We ligated both digestions with T4 ligase and transformed the products into Top10 E.coli via heat shock. We spread out on CMP-LB agar plates and incubated the plates overnight at 37°C. We analyzed obtained colonies by colony PCR and agarose gel electrophoresis. Positive colonies were inoculated in 5 ml LB with CMP and plasmids were isolated after incubating for 16 h at 37°C gaining the BioBrick BBa_K1497007. For further verification we send the samples to Eurofins for sequencing.
