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Make and run an agarose gel: Difference between revisions
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→Load samples, run gel and visualise:
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* Make a mix of sample and loading buffer (0.2 volumes loading buffer per sample - mix of food colouring for dye and glycerol for weight) for each sample you want to run, then pipette each mix into a well. Standard way is, if wells are at far end from you, load ladder at far left (well 1) and load samples to right of this. Quantity to load varies. Ladder quantity also varies, but for [http://www.taq-dna.com/rich_files/attachments/DNA_Ladders_DNA_Weight_markers_DNA_leiter/1000_bp_1kb_DNA_Ladder.pdf current] ladder, use 5ul ladder to 2ul loading buffer. Easier to see what you're doing when loading samples with something black under the box. | * Make a mix of sample and loading buffer (0.2 volumes loading buffer per sample - mix of food colouring for dye and glycerol for weight) for each sample you want to run, then pipette each mix into a well. Standard way is, if wells are at far end from you, load ladder at far left (well 1) and load samples to right of this. Quantity to load varies. Ladder quantity also varies, but for [http://www.taq-dna.com/rich_files/attachments/DNA_Ladders_DNA_Weight_markers_DNA_leiter/1000_bp_1kb_DNA_Ladder.pdf current] ladder, use 5ul ladder to 2ul loading buffer. Easier to see what you're doing when loading samples with something black under the box. | ||
* Attach electrodes to gel box, negative (black) by wells, positive (red) far end from wells, and turn on current. Power supply runs at about 75V. | * Attach electrodes to gel box, negative (black) by wells, positive (red) far end from wells, and turn on current. To check if it's working, look for bubbles coming off electrodes. Power supply runs at about 75V. | ||
* After 30-45 mins, shine UV light on the gel, look through red filter to see bands. Wear googles or glasses to protect eyes from UV, and cover skin. To see better, put something black under the box. Use a good camera, try long exposure. | * After 30-45 mins, shine UV light on the gel, look through red filter to see bands. Wear googles or glasses to protect eyes from UV, and cover skin. To see better, put something black under the box. Use a good camera, try long exposure. | ||
