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  • Gibson Assembly
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Protocols

Molecular Biology

Products and stock preparation

• Primer preparation: initial dilution of ordered primers.
• Competent Cells: prepare cells for plasmid uptake.
• Competent Cells. Protocol II (Inoue method).
• Glycerol stock: stock transformed cells at -80°C.
• Agar plate preparation: preparing agar plates for cell cultures.
• Autoclave: to sterilize solutions and glassware.

Cloning, assembly, and mutations

• Transformation: introduce foreign plasmid into competent cells.
• Plating: plate transformed cells
• Liquid cultures: when cells have grown on plates, put them in liquid cultures
• Gibson assembly.
• Linear template- TetR.
• tetR Overlap Extension PCR: induce specific mutations on tetR linear template.
• tetR Site-specific mutagenesis: induce site-specific mutations on a plasmid containing tetR.
• Agarose gel electrophoresis: DNA samples analysis (can be followed by band purification).
• Colony PCR: Analyze the plated transformed cells, to see if Gibson assembly worked
• T7 extension PCR: Put T7 promoter upstream aone gene

DNA recovery

• Plasmid preparation - Miniprep: recover plasmids from a bacterial culture.
• Gel purification: recover DNA separated by electrophoresis

Biochemistry

• IPTG test: test expression of a gene downstream from Plac

Microfluidics

• PDMS two layer device fabrication
• Master microfabrication for PDMS replica molding
• MITOMI: Protein – DNA interactions
• Chemostat cell culture
• Klenow dsDNA synthesis

Worm culture