Team:UCL/Lab AzoReducateCloning Portal

BioBrick and Primer Design

Two Azoreductase enzymes will be cloned from Bacilus subtilis (strain 168) and Pseudomonas Aeruginosa (PA01) using DNA PCR amplification.

Suitable forward and reverse primers were designed for each enzymes using the following strategy.

The protein coding sequences for each enzyme were obtained from ?.

The standard BioBrick prefix for protein coding sequences and suffix sequences used in Standard Assembly and belonging to BioBrick RFC 10 were appended to the forward and reverse primers respectively.

BioBrick CDS Prefix

5' GAATTCGCGGCCGCTTCTAG '3

BioBrick Suffix

5' TACTAGTAGCGGCCGCTGCAG '3

iGM SUBF4 : 5’-ATGTCTACAGTTTTATTTG-3’ iGM PRESUB4 : 5’-GAATTCGCGGCCGCTTCTAGATGTCTACAGTTTTATTTG-3’ iGM REVSUB:5’-TTAGAATTTTGCAGCAAGA-3’ iGM SUF REV SUB:5’-TACTAGTAGCGGCCGCTGCAGTTAGAATTTTGCAGCAAGA-3’

iGM AER4 : 5’ATGAGTAGAATTCTTGC-3’ iGM PRE AER4: 5’-GAATTCGCGGCCGCTTCTAGATGAGTAGAATTCTTGC-3’

iGM REV AER :5’-TCAGGCCGACCGCGCCAG-3’ iGM SUF REV AER: 5’-TACTAGTAGCGGCCGCTGCAGTCAGGCCGACCGCGCCAG-3’