Difference between revisions of "Team:Vilnius-Lithuania/Protocols"

Agarose Gel preparation (x%)

• To prepare agarose gels we used TopVision Agarose Tablets that are manufactured by Thermo Fisher Scientific.
• Add appropriate number of agarose tablets to the electrophoresis buffer based on the table below to prepare your desired gel percentage

Note: Use a flask that is 2 to 4 times the volume of the solution being prepared.

• Before heating soak tablets in a buffer (~ 4 minutes) until tablets completely break into fine-particle slurry. Swirl the slurry to break up any remaining particles. Important: Ensure tablets break up entirely. Heating will render non-dispersed agarose particles insoluble.
Note: Heating times are dependent on the volume of liquid and number of gel tablets to dissolve.
• Remove the flask from microwave, swirl gently to dissolve any remaining agarose particles.
• Reheat on high power for 1-2 minutes or until the solution is clear and all particles are dissolved.
• Remove the flask from the microwave oven, and gently swirl.
• Cool the solution to approximately 50-60 °C.
• Add ethidium bromide (EtBr) to a final concentration of approximately 0.2-0.5 μg/mL (usually about 2-3 μl of lab stock solution per 100 mL gel). Mix well.
• Pour the gel into a tray of required size and place in the well comb, let the gel cool and solidify for 10-15 mins at room temperature.
• This gel can now be used to run electrophoresis gels.

Note: To let the gel cool down to the required temperature of about 60 °C one can use a smaller volume container, which is not affected by heat to let a smaller volume of the melted agarose cooldown faster.