Wet lab basics

Welcome to the wet lab

These are general guidelines to help you work efficiently and to prevent you from disrupting others' experiments in the lab.

Your bench

Your bench is your area. You can pretty much expect no one to touch things that are on your bench. But that means you should also not touch other people's stuff on their benches. If you need to borrow a reagent, best to ask them in a nice way. If they're not around, Slack them. If you can't reach them, you can probably borrow it, but just be really careful with it and put it back where you found it.

Your pipettes

These are your tools - take care of them and don't use others' pipettes. Clean them with 70% ethanol regularly. Turn them up to the highest volume at night before you leave so the spring can relax. Never increase the setting higher than that.

Sterile and autoclaved materials

• Many experiments require sterile technique and tools. This is another reason to avoid borrowing things from other people’s benches – you do not want to contaminate any of their things. For this reason, it is normal practice for each person to have their own sterile water, sterile pipette tips, and sterile media. All of these are available so there should be no reason to borrow these things from other people. Once you open media, it's yours, so keep it on your bench.
• If you can, aliquot from common stocks into a separate container to avoid potentially contaminating the common stock. Things that are sterile are marked with autoclave tape. Autoclave tape has white stripes that turn black after the tape has been sterilized in the autoclave. Check the tape to make sure that whatever you think is sterile has in fact been sterilized.

Dishes and sinks

• We are lucky to have Katrina helping us with the dishwashing and dry autoclaving. We need to help Katrina by rinsing out all dirty glassware (scrub off any cell pellet residue, dried media, etc) and putting the dirty things in the large box next to the sink in room 503. Please be careful if you are working with any chemicals or products that are in any way hazardous. Make sure everything is rinsed extra well!
• Please keep the sinks clean. Don't throw any chemicals or non-soluble products in the sink (like pipette tips can go down the drain and clog the sink). If you are making gels or plates, do not pour liquids down the drain that will harden and clog the drains, e.g. agarose for DNA gels.

Autoclaving and dishwashing

The dishwasher on the 5th floor across the hall from our lab currently works - feel free to use it. The autoclave is going to be replaced. In the meantime, we use the 9th floor autoclave. If you need sterile materials, prepare them with foil and autoclave tape, put them on a lab cart in an autoclave bucket, and go up there with a labmate.

Deliveries

• When boxes arrive, please unpack them. You can let whoever's thing it is know that their thing came, or if you don't know whose thing it is, send a general message on Slack.
• Store things in the fridge, at -20C, or at -80C right away if the item requires it.
• If the delivery person asks, please sign and write the date of arrival on the packing slip. You can put packing slips on the desk near the back windows in room 503 with the other packing slips.

Common materials

You are part of a lab, so it is important to be a good lab citizen.

• If you use up some reagent or product (or even if it is close to empty) it is important to make sure more is ordered. When in doubt, let Anum know when supplies are low.
• Enzymes must be treated correctly. Everyone uses them. Enzymes need to be kept at -20° C at all times. They are stored in the freezer. You can take them out on ice or in a special blue storage block that is currently in the freezer compartment of our fridge. This block can be removed and used on your bench for several minutes and it will keep the enzymes cold. Do not leave the enzymes on your bench for more than 5 minutes. If a tube of enzyme is almost empty, please let Anum know to order a new one (we can do this through the NEB Freezer program).
• Do not vortex enzyme tubes (this will kill the enzyme).
• Only dispense enzymes with clean tips – you do not want to contaminate the enzyme tube with anything.

Common equipment

• Centrifuges
  • Anytime you centrifuge, all tubes need to be balanced. Unbalanced centrifugation is damaging to the centrifuge and can become dangerous in certain cases. Balance everything.
  • If you need it cold, keep the lid closed; otherwise, set the temperature up or turn it off and leave it open.
• NanoDrop: after use, please wipe the sample pedestal with a kimwipe wetted with deionized water, followed by a dry kimwipe.
• Shakers: if you are doing a large-scale expression and need particular temperatures or a lot of space, check with everyone else to make sure you won't be disrupting someone else's experiment.
• FPLC
  • SEC runs in particular can take a long time, so check with others to make sure everyone can schedule their run. Be considerate... some proteins are less stable, and some proteins we don't know, so collectively prioritize those.
  • Try to get your samples off as soon as your run is done.
  • Make sure to leave the buffers full, the waste beaker empty, the lines washed, the valves purged, and the fraction collector empty.
  • Save your runs with your initials and date.
  • After you use columns, equilibrate them in water (for short periods) or 20% ethanol (for longer periods).
  • Filter everything that goes on the FPLC and degas all buffers. No cell lysates.
• Gel area
  • Keep this area clean and tidy by retrieving your gels right away after you run them. You can store DNA gels short-term in a covered box in the fridge in TAE buffer, but the DNA will diffuse over time. Wipe everything down with water after use, including gel boxes and gel-related materials. If you won't be running more gels, wash out the gel boxes. Put all items away in drawers after use to leave the area open for others. Gelred is light-sensitive.
  • Chemidoc gel imager: trays are in a drawer underneath the imager, please rinse and wipe dry with a paper towel after use.
  • UV transilluminator: wipe this down after use, and use the built-in shield as well as the face-shield (hanging on the wall near the transilluminator) when cutting out bands.

Locations of certain common reagents

• All common enzymes are in the -20° C freezer in room 501.
• IPTG and TCEP are in 1 mL aliquots (1M, 400 mM) in the freezer compartment of the fridge in room 503.
• Antibiotics:
  • Ampicillin is dissolved in 70% EtOH and in the fridge. It is 100 mg/mL, which is generally 1000x concentrated.
  • Chloramphenicol is dissolved in 70% EtOH and in the fridge. It is 34mg/mL, which is generally 1000x concentrated.
  • Kanamycin is dissolved in water. There are aliquots at 30 mg/mL in the freezer compartment of the fridge in room 503. This is also generally 1000x concentrated.
• Competent cells are always kept at -80° C. Always keep the cells cold. Only take cells you are going to use.
• Bacterial plates are kept in the fridge (4° C). The sleeves of the plates are labeled. Do not remove labels from the sleeves and confuse others.

If you have questions

Just ask someone. Everyone is willing to help. It’s better to know what you’re doing and understand correct procedures than to be guessing! If you think you’ve made a mistake, contaminated a reagent or enzyme, or noticed contamination in something, please let someone know. Always better to fix problems than to try and hide them.