Experiment 6: Introduction to the Purification of Biological Molecules

Question 1

What is a typical sequence of steps for the purification of a molecule?

Answer 1

1. Cell lysis
2. Isolation of molecule of interest
3. Detection and measurement of molecule of interest

Question 2

Centrifugation relies upon _____

Answer 2

differences in solubility and/or mass and space as the basis for separating different molecules.

Question 3

Looking at an SDS-PAGE, what does a single band indicate? What about multiple bands/ a smear? What does the intensity of the band indicate?

Answer 3

A single band in an SDS-PAGE indicates that the protein collected is pure, as multiple bands or smears indicate that the protein is not pure and contaminating proteins are present in the sample that was loaded on to the gel. The intensity of the band indicates how much of the protein is present. The more intense = the more protein and vice versa.

Question 4

What are restriction enzymes?

Answer 4

Restriction enzymes are proteins that cut the sugar phosphate backbone of DNA only when a specific sequence of nucleotides is present.