Question

Question 2: EcoRI, EcoRII, and EcoRV are all restriction enzymes found in E.coli. How does E.coli prevent these enzymes from digesting its own DNA? Question 3: When performing a restriction digest it is necessary to incubate the enzymes and the DNA together to give the enzymes time to work. Given that we're using enzymes isolated from E.coli, a human symbiont, what temperature would you incubate the enzymes at, and why? Question 4: Why do DNA fragments migrate across the gel when the gel electrophoresis apparatus is running?

Answer 1

Q. 2. The restriction enzymes generally have capability of modification by methylation. By methylation, bacterial DNA will modify and hence bacteria protects its own chromosomal DNA against these restriction enzymes.

Q. 3. The enzymes of E. coli are incubated at around 37 degress celsius as this is the temperature where they perform at optimal level and it is the normal body temperature of humans.

Q. 4. DNA is negatively charged because of the Phosphate groups attached. During gel electrophoresis, electrodes are used to separate DNA particles according to their size and in it the negatively charged DNA particles moves towards the positive electrode that is the anode. Hence, the movement of DNA on gel is due to its negative charge.