Difference between revisions of "Team:TU Darmstadt/Notebook/sec1/K1602010"
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<figcaption><br><b>Figure 1</b> PCR of cadA (X). The size of the amplified product was around 1.5 kbp. DNA marker: 2-Log DNA Ladder (NEB).</figcaption> | <figcaption><br><b>Figure 1</b> PCR of cadA (X). The size of the amplified product was around 1.5 kbp. DNA marker: 2-Log DNA Ladder (NEB).</figcaption> | ||
</figure> | </figure> | ||
| + | <p align="justify"> | ||
The <I>xylC</I>-gene was synthesized by GeneArt and amplified | The <I>xylC</I>-gene was synthesized by GeneArt and amplified | ||
| − | + | by PCR using the <I>xylC</I> (FW) and <I>xylC</I> (REV) | |
oligonucleotides. The PCR product was verified by agarose gel | oligonucleotides. The PCR product was verified by agarose gel | ||
electrophoresis, digested with <i>Dpn</i>I for 120 minutes and subsequently | electrophoresis, digested with <i>Dpn</i>I for 120 minutes and subsequently | ||
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Plasmid DNA was isolated from positive clones which were verified by | Plasmid DNA was isolated from positive clones which were verified by | ||
sanger sequencing. | sanger sequencing. | ||
| + | </p> | ||
</div></html> | </div></html> | ||
Revision as of 17:10, 17 September 2015
K1602010 - D-xylonolactone lactonase xylC
Figure 1 PCR of cadA (X). The size of the amplified product was around 1.5 kbp. DNA marker: 2-Log DNA Ladder (NEB).
The xylC-gene was synthesized by GeneArt and amplified by PCR using the xylC (FW) and xylC (REV) oligonucleotides. The PCR product was verified by agarose gel electrophoresis, digested with DpnI for 120 minutes and subsequently purified. The xylC amplicon was cut with EcoRI and PstI and ligated into a pSB1C3 vector using T4-ligase. E.coli Top 10 has been transformed with the ligation product by heat shock transformation and the cells were spread out on a CMP agar plate. Clones were screened by colony PCR using VF2 and VR oligonucleotides. Plasmid DNA was isolated from positive clones which were verified by sanger sequencing.