Principles and Practice of Agarose Gel Electrophoresis

Title: Principles and Practice of Agarose Gel Electrophorsis

Objectives: To detect the size , shape and charge of the each dye solution.

Methods:

Casting the Agarose Gel

In this experiment .8% solution was used. By using a 250ml flask the buffer solution was prepared. Using the equation to make enough solution for the intire lab class the equation had to be multiplyed by five.

The contents of this equation were added to the 250ml flask and swirled to evenly distrubute it contents. Then a mark was placed on the outside of the flask to indecate the level of the solution before heating. The flask opening had perafilm placed over it so that there was little to no evaporation. The solution was then placed in the microwave and heated. The solution was then heated for one min and swriled for evenly dissolved

agarose. The agarose was then cooled, so that it was not to hot and the plate would crack. Some water was added to the solution because of there was some evaporation during heating. Once the gel had cooled , it was poured into the plate between the rubber dams. The plate was filled about half way up the comb arms. These dams are placed in the plate to prevent leaking. Then the gel was added and allowed to compeletly soiditify, which takes around 20mins.

Preparing the Gel for Electrophoresis

Once the rubber dams have been removed ( carefully) , the comb was then removed. Then the buffer was made. The buffer was made by using the equation, but also multypling it by five, for the five lab groups.

Then the chambers around the gel plate is filled with the buffer, just enough buffer to cover the gel plate in a very small amount. Then the dyes were loaded to there correct wells. Once the gels were added ( carefully) the lid was placed on the plate and system was turned on. The system ran for about 10mins. ( Hint the system is running when there are bubbles occurring

in the buffer solution.)

Results

*Look at attached page for diagram of the gel.

The results indecate that the orange dye was the smallest of the dyes. This is because it traveled the farthest from the wells. The purple and the practice dyes were the same size because they both move the same distance from the wells. The red dye is showen