Team:BCCS-Bristol/Protocols-Agarose Gel Electrophoresis



Gel preparation:

 * 1) 	For a thick gel in a small chamber, boil 50 ml 0.5x TBE in the microwave (for a thinner gel to insert only 5 µl sample in each well 40 ml are sufficient)
 * 2) 	Cool the solution until you can touch it with your hands for a longer time
 * 3) 	Add 0.5 µl ethidium bromide per 10 ml TBE and mix while avoiding air bubbles
 * 4) 	Pour the gel into a chamber with a comb that you prepared before
 * 5) 	Let the gel cool down and become solid (15-20 min)

Sample preparation:

 * 1) 	Add 10 % “Sample Loading Buffer” (BIORAD) to the sample
 * 2) 	Mix gently and spin shortly down

Loading the gel:

 * 1) 	Remove the comb and put the gel with the slide into a chamber (the wells need to be on the end with the black pole!!!)
 * 2) 	Fill the chamber with 0.5x TBE until the gel is covered
 * 3) 	Use 5 µl HyperLadderI (BIOLINE)
 * 4) 	For colony PCR, put 5 µl of each reaction in the well
 * 5) 	Close the chamber with the lid (black pole to black=negatively charged and red to red=positively charged…)
 * 6) 	Run the gel with 80-100 V

The Sample Loading Buffer contains two dyes: Bromophenol blue runs at ~300 bp and Xylene cyanol FF runs at ~4 kb.