Difference between revisions of "Team:TU Darmstadt/Notebook/sec1/K1602017"

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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>
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Revision as of 13:55, 18 September 2015

K1602017 - D-xylonic acid producing operon (T7-B0034-xylB-B0034-xylC)


https://static.igem.org/mediawiki/2015/8/8e/TU_Darmstadt_CPCR_t7-xylBC_composite.png

The BBa_K1602016 part was combined with a T7-promotor for enabling protein expression

B0034-xylB-B0034-xylC-pSB1C3 plamid was cut with XbaI and PstI and ligated into a T7-RFP-pSB1A2 plasmid which was cut with EcoRI and PstI. E. coli Top 10 cells were transformed with the ligation product. The resulting colonies were screened for positive clones by colony PCR using VF2 and VR oligonucleotides. Plasmid DNA was isolated from positive clones and verified by sanger sequencing. In addition to that cells were induced with IPTG for 12 hours at 28°C. The expressed protein was validated with SDS-PAGE.