Materials

1. Sample DNA/ Plasmid Extract

2. Ethidium bromide solution
3. 1 Tris/Borate/EDTA (TBE) buffer solution
4. RNAase
5. Dry ice or ice cubes
6. Loading dye
7. DNA marker 1 kbp
8. Agarose powder
9. Gel electrophoresis set and power supply
10.Polaroid film and photo gel set
11.Hot plate
12.Micropipette and tips (1000l, 200l, 20l )

Procedure

A. Preparing the agarose gel

1. 0.35g Agarose powder wan measured and boiled in 50 ml 1 TBE
until it is crystal clear.
2. The rack and the comb was set to solidify the gel accordingly.
3. After 30 minutes, the comb was removed gently from the gel and
gel was put into the case.
4. Enough TAE Buffer was added so that there is about 2-3 mm of
buffer over the gel.

B. Loading the gel

1. Prior loading to the gel, 1l of RNAase was added into the sample
DNA/plasmid tube and been mixed gently.
2. 10l of sample DNA/plasmid was transfer to a microcentrifuge tube
and added with about 2l of loading dye.
3. 10l of DNA marker was transfer to microcentrifuge tube and added
with about 2l of loading dye.
4. The sample and the DNA marker were loaded into the well gently.
The well was recorded accordingly.

C. Running the gel

1. The lid was place on the gel box and connecting the electrode.
2. The electrode wires was connected to the power supply, make sure
the positive (red) and negatives (black) are correctly connected.
3. The power supply was turned on to about 100V. Maximum allowed
voltage will vary depending on the size of the electrophoresis
chamber. (it should not exceed 5 volts/cm between electrodes).
4. The current is ensure to running in the correct direction by
observing the movement of the blue loading dye.
5. Then, the power run until the blue dye approaches the end of the
gel.
 6. Power was turned off and the wires was disconnect from power
supply.
7. The lid of electrophoresis chamber was removed and using the
gloves, the tray and gel was carefully removed.

D. Gel staining
1. Using gloves, the gel was removed from the casting tray and placed
into the stain dish.
2. The gel in Ethidium bromide solution was stained for 30 seconds
and destained with dH2O for 20 minutes.
3. The DNA band was visualize with UV light and the photo of the gel
was taken.
4. The DNA band samples was compared and estimated by using the
DNA markers band.