20.109(F10): TA's notes for module 3

TA Notes
Archive Fall 2009 [[Media:TA NotebookGrading.jpg| Notebook grading checklist]]

In advance of Day 1
 * Make up overnight inoculum of phage for Thursday class: 10 mL LB, 10 µL XL1-blue, 10 µL tet, 10 µL virus (from supernatant or PEG precipitated); rotate at 37ºC overnight
 * autoclave/sterilize 16 125-mL flasks (one flask per group)
 * make sure there are enough Oak Ridge tubes for each group (=3 per group)
 * make sure that there is a centrifuge available for Thursday and Friday afternoon for the PEG precipitation (Niles lab down the hall?)
 * make enough PEG solution (20% PEG-8000, 2.5 M NaCl --- this requires a long time to fully solubilize - leave on the stir plate for about an hour)

Saturday before Day 1
 * Aliquot 40 mL LB and 40 µL Tet into each eight 125-mL flasks (for Thursday class). Add 1 mL of overnight inoculum, and incubate shaking at room temperature for two days (til Monday night)

Sunday before Day 1
 * Make up overnight inoculum of phage for Friday class: 10 mL LB, 10 µL XL1-blue, 10 µL tet, 10 µL virus (from supernatant or PEG precipitated); rotate at 37ºC overnight

Monday before Day 1
 * Take off 8 125-mL flasks with 40 mL of now-saturated phage culture. Store at 4ºC (this will be for Thursday's class)
 * Aliquot 40 mL LB and 40 µL Tet into each eight 125-mL flasks. Add 1 mL of overnight inoculum, and incubate shaking at room temperature for two days (til Wednesday night)
 * check to make sure there are enough LB plates for each group to have 4 total
 * check to see that there is enough top agar for each group to have 12 mL total

Wednesday before Day 1
 * Take off 8 125-mL flasks with 40 mL of now-saturated phage culture. Store at 4ºC (this will be for Friday's class)
 * Use small (yellow-capped) tubes to make titering cells - about 1 mL is necessary for each group, so make up 6 5-mL cultures with 5 mL LB, 5 µL Kan, and a colony of ER2267. Incubate rotating overnight at 37ºC

Day 1 = Thursday
 * ice buckets (three) filled with ice for PEG precipitation
 * 40 mL phage cultures
 * titering cells (ER2267)
 * necessary chemicals (CTAB, hydrated hydrogen tetrachloroaurate, silver nitrate, ascorbic acid, and NaCl) - check their calculations before they proceed
 * Each group should make a 0.1 M solution of hexadecyltrimethylammonium bromide ("CTAB"). This is done by mixing 0.72 g with 20 ml of water in a 50 ml conical tube - CTAB will be placed in 30ºC incubator the day before to fully solubilize
 * One group will be responsible for making each of the following solutions that the class will use in the synthesis of nanowires.
 * 10 mM hydrated hydrogen tetrachloroaurate (HAuCl4•3H2O) = Mix 40 mg with 10 mL water
 * 10 mM silver nitrate (AgNO3)= Mix 17 mg with 10 mL water  -  COVER WITH FOIL!
 * 0.1 M ascorbic acid = Mix 0.18 gram with 10 mL water
 * 2.5 M NaCl = Mix 7.3 g NaCl with 50 ml water
 * store 1.5 mL aliquots with each group's sticker at 4ºC

Day after lab
 * take out titer plates from Thursday's lab and store at 4ºC

Day 2
 * quiz!
 * solutions the students made on Day 1 will be used here

Day 3
 * quiz
 * TEM grids
 * copious amounts of ethanol in both 50 mL conicals and squirt bottles
 * enough mortar/pestles for each group (=5)
 * aluminum foil

Day 4
 * make sure Belcher lab knows we're coming to use their mass balance and space, as well as someone to help with battery assembly
 * stainless steel plate and roller
 * battery demo
 * electrodes massed down in the Belcher lab and then placed in the vacuum oven to dry

Day 5
 * working with the Belcher lab to test batteries

Day 6
 * data from batteries
 * light LED

Other notes from previous TAs:
 * all antibiotic solutions are stored at 4ºC but powdered tet is in the -20ºC and powdered kan is in the 4ºC
 * taking pictures and posting them on Picasa is really fun! And it gives you a nice visual overview of what you actually did.
 * ER2267 cells are kind of picky and die after some time at 4ºC - streak out a new plate from the stock at -80ºC before beginning the module