1. (20pts) Let’s consider how restrictive enzymes work. Look the two samples of
ID: 162305 • Letter: 1
Question
1. (20pts) Let’s consider how restrictive enzymes work. Look the two samples of DNA shown below:
DNA Sample #1
5’– T C C A G T G A T C T C G A A T T C G C T A G T A A C G T T C G A T
3’– A G G T C A C T A G A G C T T A A G C G A T C A T T G C A A G C T A
DNA Sample #2
5’– T C A T C G A A T T C C T G G A A T C A G C A G A A T T C G C A T A
3’– A G T A G C T T A A G G A C C T T A G T C G T C T T A A G C G T A T
If both samples are treated with the restriction enzyme EcoRI [recognition sequence G’AATTC] then indicate the number of fragments and the size of each fragment from each sample of DNA. List the fragment sizes in order of largest to smallest.
Sample # 1 Sample # 2
# of fragments:________ # of fragments:________
Sizes of fragments (largeà small): Sizes of fragments (largeà small):
(20pts) Read the protocol for Lab 8B.
What is supercoiled DNA
Why would it migrate further into an agarose gel if it’s uncut plasmid DNA?
What causes this type of DNA to form?
Is this likely to occur to our isolated plasmid DNA?
Explanation / Answer
1. Sample 1: 2 fragments
Fragment 1: 13 base pairs (5' to 3')
Fragment 2: 21 base pairs (5' to 3')
Sample 2: 3 fragments
Fragment 1: 6 base pairs (5' to 3')
Fragment 2: 18 base pairs (5' to 3')
Fragment 3: 10 base pairs (5' to 3')
2. Supercoiled DNA: It is the type of DNA which is under or over-coiled. A DNA with unusual topology can be called a supercoiled DNA. Against an uncut plasmid, it would migrate over a lesser distance, due to more complex topology, as it would make passing through the agar pores difficult. DNA like these are formed under external circumstances related to transcriptional control and available area inside the nucleus. It should not apply to an isolated plasmid DNA as it is in absence of external factors.
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