21 June 2008
Gel Electrophoresis
- Gel Electrophoresis is primarily used to determine the length of a fragment of DNA. DNA Ladders are run beside experimental fragments as points of reference. A DNA Ladder contains all different lengths of DNA that when run out on a gel, separate to reveal the number of base pairs in each gene.
- Prepare the casting gel: 1% agarose gel - 1) 0.5 g agarose 2) 50 mL TBE + SYBR safe.
- Gel must be submerged completely with 300-500 mL (large gel tray) .5X TBE + SYBR safe buffer.
- Put the gel, WITHOUT the lid, in the microwave on HIGH for 1 minute.
- Cool the bottle of gel under water.
- Pour into the castings with rubber stoppers.
- The gel takes approximately 20 minutes to set. Once gel is set make sure to remove the rubber stopprs.
- DNA has a negative charge. The wells of the gel are aligned with on the negative side (Cathode). DNA will move away from the Cathode to the Anode (+).
- The gel acts as a sieve for fragements as small as 100 base pairs and as large as 50,000 base pairs. 0.3% large fragments - 2% large fragments.
- When complete, pour TBE buffer back into a container. The buffer can be used up to 8 times.
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