9. Lab Rules
• No food & drink in the lab
• Gloves & aprons at all times
• Wash/gel your hands when you leave the
lab
• Aargh spills!
• No running, diving or bombing
Sunday, 10 February 13
10. Aseptic Technique
• Keep air exposure to a minimum.
• Don’t cross-contaminate.
Sunday, 10 February 13
11. Micropipetting
• Two different types
• Set the volume by turning the dial
• Suck!
• Squirt!
• Remember:Always use a fresh tip.
Sunday, 10 February 13
13. 1.
• Iron Microbeads! (In tube 1)
• Wash them… 50µl of washing buffer. Mix.
• Clean them… pull the beads to one side
with the magnet; extract the liquid.
• Wash and clean them again…
Sunday, 10 February 13
14. 2.
• Ligation! Glues the first “part” to the
beads.
• Add 7µl of ligation mix (tube with the spot
on) and mix.
• Transfer this to the tube containing our
first part (Tube 2)
• Wait 5-10 minutes…
Sunday, 10 February 13
15. 3.
• Wash them… 50µl of washing buffer. Mix.
• Clean them… pull them to one side with
the magnet; extract the liquid.
• Wash and clean them again…
Sunday, 10 February 13
16. 4.
• Ligation again! Glue the final two parts to our beads.
• Add 7µl of ligation mix (tube with the spot on) and mix.
• Transfer this to the tube containing our second part
(Tube 3) and mix.
• Transfer this to the tube containing our first part (Tube
4) and mix some more.
• Wait 5-10 minutes…
Sunday, 10 February 13
17. 5.
• Wash them… 50µl of washing buffer. Mix.
• Clean them… pull them to one side with
the magnet; extract the liquid.
• Wash and clean them again…
Sunday, 10 February 13
18. 5.
• Elution! Gets rid of the beads
• Add 20µl of elution buffer and mix
• Pull the beads to one side with the magnet
• Extract the finished part in liquid form; put
it into one of the empty tubes.
Sunday, 10 February 13
20. Modify that e.coli!
• Put 100µl of Transformation buffer in an empty tube; put
the tube on ice for 5 minutes, then…
• Take a loop. Grab some e.coli and add it to the
transformation buffer – “twiddle vigorously”.Wait for 10
minutes…
• Add 40µl of your creation and mix.
• Bring your tube to the front!
Sunday, 10 February 13
21. Modify that e.coli!
(Pt. 2)
• “Heat shock” our e.coli for exactly 30
seconds in the water bath (42ºc)
• Put them back on ice for 1-2 minutes
• Add 100µl of recovery broth, keep them
warm in your hand until…
Sunday, 10 February 13
22. Modify that e.coli!
(Pt. 3)
• Put 100µl of your modified e.coli in a petri
dish.
• Remove the spreader from the alcohol and
flame it (so there’s no alcohol left)
• Spread the e.coli around your plate
• …and that’s it! Bring your plate to the front
and put it in the incubator.
Sunday, 10 February 13
