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SUBMARINE GEL ELECTROPHORESIS

Requirements :
1.Submarine gel system
2.Mini power pack
3.UV Transilluminator.
4.DNA samples, standard marker

Solutions:
1. SB Buffer:
To prepare 250 ml, weigh
Sodium hydroxide - 0.1 9
Boric acid - 2.0g
Dis.water make up to - 250ml(pH 8.0)

2.6X gel loading buffer (40% sucrose, 0.025%BPB) To


prepare 1 ml
Sucrose - 0.040g

BPB - trace
Dis.water make up to - 1 ml

3.Ethidium Bromide (5μg/ml)


To prepare 1 ml stock solution
Ethidium bromide - 5 mg
Double dis .water up to - 1 ml

4. Agarose ( 1 %) (2%) 0.2g 10ml


Agarose 0.1 9
SB buffer 10 ml
PROCEDURE:

1. Clean the gel platform and comb seal the open ends with cellophane tape.

2. Keep the Agarose solution (10ml) in a boiling water bath and dissolve the
agarose.

3. Cool down the agarose solution temperature to 56C.

4. Add 1μl Ethidium bromide to the Agarose solution, mix gently and Pour on
to the gel platform gently.

5. Place the comb in the notch, and allow the gel to set.

6. Place the gel platform with comb in the submarine unit.

7. Pour 165 ml of SB buffer 2-3 mm above the surface of the gel.

8. Add 2ul of 6x gel loading buffer to 10μl of sample.

9. Keep it in boiling water bath for 3 minutes.

10. Remove the comb slowly from the gel and inject samples slowly in to the
wells.

11. Continue the run till the BPB tracking dye reaches the anodic end.

12. Observe the bands in a UV transilluminator.