Team:TU Darmstadt/Notebook/sec1/K1602019

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K1602019 - B0034-Humicola insolens cutinase (B0034-HIC)

K1602018 was used as template for a PCR with oligonucleotides His-HIC and HIC_rev. The PCR product was digested with DpnI, purified and used as template for a second PCR with oligonucleotides Prefix-HIC and HIC_rev. The construct was verified by gel electrophoresis and digested with XbaI and PstI and ligated into B0034-pSB1A2. Competent E. coli Top 10 were transformed via heat shock. Positive colonies were inoculated and the plasmids were extracted. The plasmids were digested with EcoRI and PstI and ligated into pSB1C3. Competent E. coli Top 10 cells were transformed via heat shock and the plasmids were extracted.