Difference between revisions of "Team:TU Darmstadt/Notebook/sec1/K1602030"
| Line 4: | Line 4: | ||
<html><div class="contentSection"> | <html><div class="contentSection"> | ||
| − | <a href=" https://2015.igem.org/Team:TU_Darmstadt/Notebook/sec1/K1602027" title="Opens internal link in current window" class="internal link">K1602027</a> was used as template for PCR with oligonucleotides VF2 and sc-Si4-REV1. After digesting the PCR product using <i>Dpn</i>I and purification the silica-tag Si4 was connected to the scaffold by another PCR using the oligonucleotides VF2 and sc-Si4-REV2. A gel extraction was performed. The purified PCR product was then digested with <i>EcoR</i>I and <i>Pst</i>I. The construct was ligated into pSB1C3 and transformed into E. coli Top 10 via heat shock. Colonies were screened by colony PCR with VF2 and VR. | + | <a href=" https://2015.igem.org/Team:TU_Darmstadt/Notebook/sec1/K1602027" title="Opens internal link in current window" class="internal link">K1602027</a> was used as template for PCR with oligonucleotides VF2 and sc-Si4-REV1. After digesting the PCR product using <i>Dpn</i>I and purification the silica-tag Si4 was connected to the scaffold by another PCR using the oligonucleotides VF2 and sc-Si4-REV2. A gel extraction was performed. The purified PCR product was then digested with <i>EcoR</i>I and <i>Pst</i>I. The construct was ligated into pSB1C3 and transformed into <em>E. coli</em> Top 10 via heat shock. Colonies were screened by colony PCR with VF2 and VR. |
</div></html> | </div></html> | ||
Latest revision as of 07:23, 18 September 2015
K1602030 – HistidinTag-Scaffold-SilicaTag4 (His-Scaffold-Si4)
K1602027 was used as template for PCR with oligonucleotides VF2 and sc-Si4-REV1. After digesting the PCR product using DpnI and purification the silica-tag Si4 was connected to the scaffold by another PCR using the oligonucleotides VF2 and sc-Si4-REV2. A gel extraction was performed. The purified PCR product was then digested with EcoRI and PstI. The construct was ligated into pSB1C3 and transformed into E. coli Top 10 via heat shock. Colonies were screened by colony PCR with VF2 and VR.