Experiments

1. Add tap water until the drainboard at the bottom of the can is slightly submerged.
2. Place 1.5 mL tubes in a sterile bag and place it on top of the drainboard.
3. Select Sterilize and Keep Warm and start at 121 ℃ and 15 minutes.
4. Wait approximately 1 hour and 15 minutes.

1. Prepare agarose gel (1%).
   1. Add 400 mL of 1x TAE to an Erlenmeyer flask.
   2. Add 4 g of agarose.
   3. Heat the solution by microwave until it boils, while stirring appropriately.
   4. After the solution has cooled to 50 ℃, add 16 μL of Midori Green solution.
   5. Pour the solution into the plate and leave for one hour.
   6. Store the gel on a kimtowel moistened with nuclease free water.
2. Place the prepared gel on the gel bed of the migration tank.
3. Pour buffer solution to a level of about 3 mm above the surface of the gel.
4. Apply 10 µL of the sample to the wells.
5. Start migration.
6. After preparing the dye in a sealable container, remove the gel tray from the migration tank and submerge it in the stain solution.
7. After allowing it to stand for the required amount of time, remove the gel and place it on the transilluminator for observation.

1. Check that the cleaning water bottle contains Dw and that the waste liquid tank is not full, and then clean the probe.
2. Perform microchip cleaning.
3. Register the samples on the capillary electrophoresis device MultiNA and set the wells to contain the samples.
4. Dilute a solution of the fluorescent dye (diluted 100 times with TE) 100 times with DNA-500 (gel) to create a separation buffer.
5. Place the calculated amount of the DNA marker displayed on MultiNA into a 0.5 mL tube.
6. Cut the 8-tube strips into 2-tube strips, and add 15 µL of the size ladder to one tube.
7. Set the cleaning solution, separation buffer, DNA marker, and size ladder in the designated positions.
8. Dispense 10 µL of the electrophoresis samples into 8-tube strips and place them in the wells set in step 3.
9. Start analysis.

Ref. Shimadzu Corporation. (n.d.). Microchip Electrophoresis System for DNA/RNA Analysis MCE-202 MultiNA Instruction Manual. http://mor.niboch.nsc.ru/public/MBA_Course/References/Electrophoresis/devices/Shimadzu%20MultiNA%20manual.292-28464D_SWmanualMCE-202(E).pdf

1. Blank with 1 µL of TE.
2. Run 1 µL of samples.
3. Rinse with 1 µL of TE after each sample.

1. Set up the QuantStudio 3 Real-Time PCR System. Modify experiment settings as needed.
2. Load a 96well plate in the instrument.
3. Start the run.

1x HCR Buffer Components

20 mM Tris-HCl (pH. 8.0)
140 mM NaCl
5 mM KCl

1x LT Buffer Components

10 mM KCl aq.
10 mM Tris-HCl
18 mM MgSO₄ aq.
10 mM (NH₄)₂SO₄ aq.
pH8.7 at 25℃

1x UTokyo Buffer Components

60 mM KCl aq.
10 mM MgCl₂ aq.
10 nM CoCl₂ aq.
5 mM HEPES-NaOH
pH 7.7 at 25 ℃