Day 3 Exp 1

Question 1

what is the purpose of the alkaline lysis method and spin column

Answer 1

we are trying to separate the DNA from the macromolecules

Question 2

how will we use restriction enzymes in this experiment

Answer 2

we will use them to cut the DNA at specific sites and see how the fragments have the same or different patterns from eachother

Question 3

what might cause the DNA sequences to change and be cut differently

Answer 3

mutations

Question 4

if the DNA sequence was changed to have an additional cur site, what might happen to the fragment

Answer 4

it would be cut into two instead of one

Question 5

if the sequence was changed so that the bases no longer matched the recognition sequence of the enzyme what would happen to the fragment

Answer 5

it would not longer be cut

Question 6

if the insert has additional cut site within its own sequence, what will happen to the fragment

Answer 6

it will be different from other clones that do not

Question 7

what are chaotrophic Reagents?

Answer 7

they are in the lysis buffer and disrupt the interaction of DNA and water, allowing it to bind to the silica membrane

Question 8

how do chaotropic reagents work

Answer 8

they disrupt hydrogen bonding between DNA and water and they also interfere with hydrophobic interactions and proper structure of biological molecules

Question 9

what do chaotrophic reagents do to proteins?

Answer 9

they denature them

Question 10

What step of the miniprep procedure gets rid of large molecules

Answer 10

The addition of an Alkaline lysis buffer with chaotropic reagents allows the DNA to bind to the silica. The neutralization and precipitation of unwanted cellular components allow us to form a pellet with the debris.

after centrifugation, we can then extract the supernatant and leave the cell debris behind in the pellet