User:Jarle Pahr/Checklists

Checklists for lab procedures:

Start of day:

• Wipe down lab bench with ethanol.

General experiment checklist:

• Prepare all materials the day before. Ensure that all necessary items are available/reserved and ready to use, including flasks/bottles. Tubes, pipette tips, etc.
• If possible, perform a dry run of the experiment beforehand to. This may include pipetting of water to simulate samples, if pipetting operations might need planning.

Fluorescence experiments:

• All necessary strains are inoculated, including controls.
• Appropriate controls are included.
• "Gain" setting is set to automatic or manual, as appropriate.
• Instrument is reserved.

Transformation:

• Supercompetent cells are available (remove from freezer 20 min before use, to avoid waiting for cells to thaw).
• Enough plates, with right antibiotic, are available. Remove from cold storage, place in incubator for pre-heating.
• Waterbath for heat shock is at right temperature (42 C).
• Ethanol for sterilization of glass spreader is available. (use absoulute ethanol).

Before heat shock: Place plates in incubator for pre-heating.

Restriction digest:

• Needed restriction enzymes are available.
• Thaw buffer
• Waterbath at 37 C is available. (or otherwise, thermomixer, or otherwise, shaking incubator)

Ligation:

• Ligation buffer is available.
• Ligase is available.
• Correct ligase is used (T4 DNA ligase)
• If possible include positive control to make sure the ligase is functional.
• If possible, include negative control of bacbone only/insert only, as appropriate.

Pipetting:

• Release plunger slowly to avoid air bubbles forming at the tip.
• When pipetting small volumes (enzyme reactions, etc.), confirm both that the right volume is drawn into and expelled from the pipette.

Preparation of agar plates:

• Magnetic stirrers are available
• Antibiotic is available. Thaw antibiotic.

Preparing supercompetent cells:

• Centrifuge is cooled. Chill tube holders of the correct size by placing them in the centrifuge in advance.
• Spectrophotometer and cuvettes for measuring OD are available
• When using spectrophotometer: Correct wavelength is selected (600 nm)
• For snap freezing: Dry ice or liquid nitrogen is available.
• Freezer box is available
• Sterilize tubes for cell aliquots are available.
• Freezer box is marked.

Electrophoresis:

• Imaging system is operational
• After imaging: Save image as image file. Copy file(s) to personal area.
• Remove gel from imaging apparatus.
• Copy image to document, write observations.

Gel extraction:

• The correct band is cut.

Measuring OD:

• Instrument is set to correct wavelength (typically 600 nm)
• Cultures are re-placed in incubator immediately after sampling.
• Enough liquid in cuvette
• Cuvette is inserted in correct orientation

Miniprep:

• Centrifuge is available
• Before centrifuging, measure OD600 is possible, for records on relation between culture properties and DNA yield.
• After measuring DNA concentration, record statistics (culture volume used, OD600, yield, etc.) for use in procedure optimization.

Use of instruments, general:

• After end of run/experiment, copy data to personal area.
• Print paper of copy of raw or processed data, place in ring binder or lab journal.

Inoculating cultures:

• A suitable incubator and tube rack / holder for the culture(s) is free.

Receiving plasmid samples for use:

• As soon as possible, verify the sequence (entirely, or critical parts only) for yourself
• Make a new glycerol stock for your personal use.

End of day:

• No samples are left on bench (that shouldn't be there).
• Update sample records
• Wipe down lab bench with ethanol.