Gel electrophoresis

Polyacrylamide gels?

(vertical gels used to separate proteins) create an even smaller matrix and are used to separate proteins by size. Prior to running proteins we denature them, so that they are all negatively charged

How does gel electrophoresis separate molecules?

Agarose gels create a matrix that molecules move through. Small molecules move faster through the matrix. Molecules move through the matrix through an electric current, since DNA is negatively charged, all DNA moves to the positive electrode. Polyacrylamide gels (vertical gels used to separate proteins) create an even smaller matrix and are used to separate proteins by size. Prior to running proteins we denature them, so that they are all negatively charged.

By which electrode do you put the wells in DNA gel electrophoresis?

Since DNA is negatively charged, you place the wells by the negative electrode so that the DNA will move toward the positive electrode.

Do smaller or larger molecules move faster through the gel?

Small molecules move faster, large molecules move slower

How does the percentage of agarose affect the rate of movement/migration through the gel?

The more percentage of agarose present, the thicker the gel and the easier smaller pieces of DNA can be separated. The less percentage of agarose present the thinner the gel and the better the gel will separate very large pieces of DNA.

What is the purpose of a standard curve?

The purpose is to create a way to find the size of an unknown

What is the purpose of gel electrophoresis?

To separate DNA (or other molecules, such as proteins) by size and charge.

How do you make DNA gels?

You determine the percentage of agarose needed in the gel, and use that same number in grams, and the balance in buffer. For example, if you want a 2% gel, use 2 grams of agarose powder and 98 ml of buffer. If you want 0.5% gel you use 0.5 grams of agarose powder and 99.5 ml of buffer.

How do you use the standard curve?

You use the distance migrated of the unknown and find the corresponding weight/size on the standard curve.

How do you create a standard curve?

You use the known weights or sizes of the molecules, as well as the distance migrated of each band to create a line that serves as a standard curve.

Agarose gels?

create a matrix that molecules move through. Small molecules move faster through the matrix. Molecules move through the matrix through an electric current, since DNA is negatively charged, all DNA moves to the positive electrode.