Team:TU Darmstadt/Notebook/sec1/K1602016

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K1602016 - D-xylose dehydratase plus D-xylonolacton lactonase with strong RBS (B0034-xylB-B0034-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 BBa_K1602000 part and the BBa_K1602001 part were combined

Firstly the B0034-xylB composite was digested with EcoRI and SpeI while the B0034-xylC composite was digested with XbaI and PstI. They both were taken from a pSB1A2 plasmid. The restriction products were dephosphorylated and ligated in one step into a pSB1C3 vector 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.