Topic 2: Meselson and Stahl (DNA replication)

Question 1

In the lab, we can grow bacteria in a nutrient solution containing ammonium ions (NH4+) or nitrate ions (NO32-)
The bacteria use the _______ to make DNA _____.

Then they use the DNA _____ to make _______________.

Then they use the _______________ to make ______.

So the nitrogen from the nutrient solution becomes incorporated into the bacteria’s DNA __________.

Answer 1

nitrogen

bases

bases

mononucleotides

mononucleotides

DNA

molecules

Question 2

Meselson and Stahl used 2 isotopes of nitrogen in their experiments.

What 2 isotopes were they?

Answer 2

1. 14N - lighter isotope of nitrogen (common)

2. 15N - heavier isotope of nitrogen

Question 3

If bacteria are grown in a nutrient solution containing ammonium ions (14NH4+ ) where the N is ‘light’ (14N) the DNA will contain 14N (light nitrogen), which makes the DNA _______.

If the DNA is extracted from bacteria and __________ (spun at high speed), as all the DNA is ‘_____’, a band of DNA forms at the _____ of the tube.

Answer 3

lighter (less dense)

centrifuged

light

top

Question 4

If bacteria are grown in a nutrient solution containing 15NH4+ ions the DNA will contain 15N (heavy nitrogen) - the extra neutron in the nucleus makes DNA ________.

If the DNA is extracted from bacteria and ___________ (spun at high speed), as all the DNA is ‘_____’, the band of DNA forms at the ________ of the tube.

Answer 4

heavier (more dense)

centrifuged

heavy

bottom

Question 5

(heavy and light nitrogen experiment)
Key Summary: What happens when DNA is centrifuged in a density gradient?
(3 points)

Answer 5

1. Heavy DNA (made from 15N) forms a band at the bottom
2. Light DNA (made from 14N) forms a band at the top
3. {Medium/intermediate} DNA (made from a mixture of 14N and 15N) forms a band in the middle of the tube

Question 6

Name the 3 theories (that Meselson and Stahl tested) about how DNA replicated

Answer 6

1. Fragmentary
2. semi-conservative
3. conservative

Question 7

Describe fragmentary replication

3 steps

Answer 7

1. The original parent DNA molecule which needs to replicate breaks down into mononucleotides (fragments)
2. The two new DNA molecules are formed from a mixture of mononucleotides from the original parent DNA molecule and the new mononucleotides made by a cell.
3. The two molecules are identical.

Question 8

Describe semiconservative replication

4 steps

Answer 8

1. The original parent DNA molecule which needs to replicate separates into two strands
2. Two new DNA strands are formed from the new mononucleotides, one alongside each of the strands from the original DNA
3. each new DNA molecule contains one strand from the original DNA and one new strand
4. The two molecules are identical

Question 9

Describe conservative replication

2 steps

Answer 9

1. The original parent DNA which needs to replicate stays intact
2. A new DNA molecule is made from new mononucleotides, which is identical to the original molecule

Question 10

Key Summary: What were the results of the semiconservative theory of DNA replication experiment?
(1 point - after 1 cycle of replication)
(3 points - after 2 cycles of replication)
(2 points - after 3 cycles of replication)

Answer 10

After 1 cycle of replication:
1. {ONE/single} band of {medium/intermediate} DNA in the middle of the tube

After 2 cycles of replication:

1. TWO bands – each 50% - bands same width
2. one {medium/intermediate} DNA (in the middle of the tube)
3. one light DNA (at the top of the tube)

After 3 cycles of replication:

1. one wider band (wider than the bands formed after 2 cycles) of light DNA at the top of the tube
2. one narrower band (narrower than the bands formed after 2 cycles) of medium DNA in the middle of the tube.

Question 11

Key Summary: Describe the method & evidence for accepting the semi-conservative theory
(Meselson and Stahl DNA replication experiment)
(1 point stating aim of experiment + 5 steps)

Answer 11

1. Experiments aimed to distinguish between 3 types of replication - conservative, fragmentary, semi-conservative.
2. Bacterial cells were grown for many generations in a growth medium with heavy nitrogen (15N) - (only a single band of heavy DNA would be observed if centrifuged)
3. The bacterial cells were then transferred to a growth medium with light nitrogen (14N) and allowed to divide TWICE
4. DNA was extracted and separated by centrifugation after ONE AND TWO CELL {CYCLES/divisions}
5. DNA extracted after one round of replication shows up as A SINGLE BAND OF INTERMEDIATE DNA which contained one heavy strand and one light strand.
6. DNA extracted after two rounds of replication showed half the DNA gave one band of intermediate DNA and half gave one band of light DNA, proving the semi-conservative theory.

Question 12

Key Summary: describe the method & evidence for refuting the fragmentary theory of DNA replication
(Meselson and Stahl DNA replication experiment)
(1 point stating aim of experiment + 5 steps)

Answer 12

1. Experiments aimed to distinguish between 3 types of replication - conservative, fragmentary, semi-conservative.
2. Bacterial cells were grown for many generations in a growth medium with heavy nitrogen (15N) - only a single band of heavy DNA was observed
3. The bacterial cells were then transferred to a growth medium with light nitrogen (14N) and allowed to divide TWICE
4. DNA is extracted and separated by centrifugation after ONE AND TWO CELL {CYCLES/divisions}
5. For the fragmentary theory to be accepted, after every round of replication an intermediate band of DNA was expected
6. After two rounds of replication one intermediate band and one light band were observed, refuting the fragmentary theory

Question 13

Key Summary: Describe the method & evidence for refuting the conservative theory of DNA replication
(Meselson and Stahl DNA replication experiment)
(1 point stating aim of experiment + 5 steps)

Answer 13

1. Experiments aimed to distinguish between 3 types of replication - conservative, fragmentary, semi-conservative.
2. Bacterial cells were grown for many generations in a growth medium with heavy nitrogen (15N) - only a single band of heavy DNA was observed
3. The bacterial cells were then transferred to a growth medium with light nitrogen (14N) and allowed to divide ONCE
4. DNA is extracted and separated by centrifugation after ONE CELL {CYCLE/division}
5. For the conservative theory to be accepted, after one round of replication one band of light DNA and one band of heavy DNA would be expected
6. After one round of replication only one intermediate DNA band was observed – refuting the conservative theory