Salk Institute Mobile Lab Gel Electrophoresis Teacher Instructions BioRad Set Up 8 groups Grossmont Kit

Salk Institute Mobile Lab Timeline Prepare Gels: Up to a week in advance Class lab: 1-3 days –Teach students to pipette –Load and run gels –Teach electrophoresis theory –Analyze gels Gels must be analyzed no later than next day after running (stored in refrigerator overnight)

Salk Institute Mobile Lab Prepare 1X TAE Buffer for making gels Measure 28ml of 50X TAE Buffer stock solution into the 50ml conical TAE Buffer measuring tube

Salk Institute Mobile Lab Pour the 28ml of 50X TAE Buffer stock solution into the 2L TAE Buffer mixing bottle Prepare 1X TAE Buffer for making gels

Salk Institute Mobile Lab Fill 2L TAE Buffer mixing bottle to the 1400ml line with water (tap or distilled) You might need to repeat this as necessary for your number of classes – this bottle should prepare enough 1X TAE Buffer for 6 classes worth of gels Prepare 1X TAE Buffer for making gels

Salk Institute Mobile Lab Measure 3 level scoops of agar powder into each glass agar mixing bottle –Each scoop is 1/4 tsp –This will equal ~1.5g of powder per bottle You’ll need to make 1 bottle per class Prepare Agar for Gels

Salk Institute Mobile Lab Fill each bottle to the 200ml mark with your prepared 1X TAE Buffer solution –Bottles have been pre- checked for calibration Cap tightly and shake to mix Prepare Agar for Gels

Salk Institute Mobile Lab Loosen caps slightly and place bottles in your microwave Set microwave for 1-2 minutes per bottle (less is better - you can always add more time!) Allow agar solution to come to a boil - stop microwave once a good boil starts Prepare Agar for Gels

Salk Institute Mobile Lab Carefully remove the HOT bottle from the microwave and swirl - be careful of steam escaping from the loose caps! Prepare gels

Salk Institute Mobile Lab Check that agar has fully dissolved or, if re-melting solidified agar, that it has all melted back into solution Allow Agar to cool until you can just stand holding the bottle with your bare hand Prepare Agar for Gels

Salk Institute Mobile Lab Prepare Gel Casting Trays While Agar is cooling use blue lab tape to carefully seal the end of each tray Place 2 combs into slots on each tray (top and middle)

Salk Institute Mobile Lab Carefully pour warm agar solution into the assembled gel casting trays (make sure agar is still fully melted) Fill each tray approximately.75cm 1 bottle should fill 4 trays Pour Gels

Salk Institute Mobile Lab After gels have solidified, remove the combs and tape Carefully slide gel out of the tray onto a “patty pac” paper 1 gel per paper Re-tape tray to pour for next class set How to store prepared gels

Salk Institute Mobile Lab Place 2 papers with 1 gel each side by side in a gel storage container Make sure paper edges are free How to store prepared gels

Salk Institute Mobile Lab Stack second layer of gels in storage container and place container in fridge for up to a week 1 container per class! –(4 full size gels) How to store prepared gels

Salk Institute Mobile Lab When you are ready to have students use gels simply carefully lift paper with gels out of the storage container Carefully use spatula to cut each gel in half (~1cm above middle wells) Setting up prepared gels for class

Salk Institute Mobile Lab Place each gel half onto flat tray for each student group –Put a patty pac under if too hard to see wells Try to keep gels and trays low and level to prevent accidental tearing of the gel Setting up prepared gels for class

Salk Institute Mobile Lab Prepare 1X TAE Buffer solution as before: –Pour the 28ml of 50X TAE Buffer stock solution into the 2L TAE Buffer mixing bottle –Fill 2L TAE Buffer mixing bottle to the 1400ml line with water (tap or distilled) Pour 1X TAE into electrophoresis boxes stopping about 1cm below MAX line Running gels

Salk Institute Mobile Lab After students have loaded their gels carefully slide each gel half back into each gel tray (in same direction) and place into the electrophoresis boxes –Keep track of which groups’ gels are where! Make sure the well sides of the gels are on the BLACK electrode side –Back to Black, RUN to RED Running gels

Salk Institute Mobile Lab Top off each box as needed with TAE to bring level to MAX line Place lid on each electrophoresis box making sure that the black electrode is at the well end of the gels Running gels

Salk Institute Mobile Lab Connect the electrodes from each box to the power supply and turn on the power by the switch in the back Make sure the power supply is set on volts and adjust the voltage to 200* When you are ready to start the run simply press the button in the center –Watch for bubbles in the electrophoresis boxes! Running gels *may need to lower voltage

Salk Institute Mobile Lab When the run is complete (colors have separated) turn off the power by pressing the button on the left Remove the lids from the electrophoresis boxes After Gel Run

Salk Institute Mobile Lab Carefully remove gel trays from the box and depending on time: –Place each ½ gel on a flat tray on a patty pac and give back to groups to analyze –OR place each ½ gel on a labeled patty pac and store back in storage container in refrigerator until next class meeting and then distribute on flat trays WARNING - WET GELS ARE VERY SLIPPERY!! After Gel Run

Salk Institute Mobile Lab Next period and so on… You can prepare per 2 gels for distribution while per 1 gels are running and so on… Running TAE buffer is good for all classes – no need to replace unless it gets too hot

Salk Institute Mobile Lab Clean up At end of day used buffer can just be flushed down sink –Rinse boxes and let air dry Used gels can be placed in general trash