Mutagenesis of protein A

Paweł

1. Isolation of plasmid DNA from overnight cultures (it was really hard work...).
2. Control digest of isolated plasmids with KpnI and BamHI (if mutagenesis really occured, ~550bp band should be visible).
3. Results of digest:
• ADelL+KpnI and ADelP primers mutagenesis:
• pACYClac+ompA-ΔA-omega: 0 positive, 5 negative
• pACYClac+ompa-ΔA-alpha: 3 positive, 7 negative
• pACYClac+OmpA-omega-ΔA: 1 positive, 2 negative
• AMutL+KpnI and AMutP mutagenesis:
• pACYClac+ompA-ΔA-omega: 4 positive, 16 negative
• pACYClac+ompa-ΔA-alpha: 4 positive, 8 negative
• pACYClac+OmpA-omega-ΔA: 2 positive, 15 negative
4. Plasmids which gave positive digest results prepared for sequencing (one sample from each variant which gave positive result).