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Team:TU-Delft/Protocol2
From 2013.igem.org
Protocols
Our project deals with E.coli cells which sense Auto-inducing peptides (AIPs) from the Staphylococcus aureus and starts producing Antimicrobial peptides in order to kill the Staphylococcus aureus. Different protocols used during the project are described below.
- Transforming Parts from Distribution kit
- Growing the Single Colonies from the Agar Plates
- Making glycerol stocks
- Plasmid Purification Protocol
- Restriction digestion
- Ligation
- Gel Extraction Procedure
- PCR Purification
- Tricine Gels
- General Peptide Production
- SUMO cleavage
- Lysis Protocol
- AIP Sensing Protocol
- Gene Design
- Primer Design
Growing the Single Colonies from the Agar Plates
Requirements:
- LB Broth
- Pipettes
- Antibiotics
- Sterile flasks
Procedure:
- Prepare flask with LB Broth in it. Add the appropriate antibiotic needed.
- Select the single colony using the pipette tip from the Agar plate, which contains the bacterial cells.
- Inoculate the LB broth with the bacterial cells.
- Grow the culture for 14-16 hours.
Next day prepare glycerol stocks and carry out a miniprep protocol to extract the desired plasmid from the bacterial cells.
