DNA Replication

Learning objectives Distinguish between DNA replication, transcription and translation Define heavy nitrogen Distinguish between conservative and semi- conservative hypothesis Conclude the correct hypothesis from the Meselson- Stahl experiment

Matthew Meselson & Franklin Stahl 1958

Conservative or semi-conservative?

How it worked . . . . They knew from the structure of DNA that it contained nitrogen. They grew bacteria (E.Coli) in solutions which contained only ‘heavy’ nitrogen (15N) After many generations, all the DNA in the bacteria was ‘heavy’ and it contain no ‘normal’ nitrogen (14N). The DNA was extracted and centrifuged in a solution of 6M caesium chloride CsCl.

Matthew Meselson & Franklin Stahl 1958 The E.Coli were then kept in 15N for many generations and the DNA centrifuged in 6M CsCl Matthew Meselson & Franklin Stahl 1958

Centrifugation in 6M CsCl Increasing concentration ‘Heavy’ DNA is found near the bottom of the tube

The E.Coli were then kept in 14N for a generation and the DNA centrifuged in 6M CsCl

Centrifugation in 6M CsCl Increasing concentration ‘Medium’ DNA is found higher in the tube

Matthew Meselson & Franklin Stahl 1958 The E.Coli were then kept in 14N for a second generation and the DNA centrifuged in 6M CsCl Matthew Meselson & Franklin Stahl 1958

Centrifugation in 6M CsCl Increasing concentration ‘Light’ DNA is found higher in the tube due to its lower density ‘Medium’ DNA was still to be found here but it was a lighter band indicating that there was less of it

Drawing Conclusions

Matthew Meselson & Franklin Stahl 1958 Generation 0 (DNA from bacteria that had been multiplying in 15N for many generations) Increasing concentration 14N 14N 14N 15N 15N 15N ‘Heavy’ DNA is found near the bottom of the tube Matthew Meselson & Franklin Stahl 1958

Generation 1 Increasing concentration 14N 14N ‘Medium’ DNA is found near the bottom of the tube

Matthew Meselson & Franklin Stahl 1958 Generation 2 Increasing concentration 14N 14N 14N 15N 15N 15N ‘Light’ DNA is found higher in the tube due to its lower density ‘Medium’ DNA was still to be found here but it was a lighter band indicating that there was less of it - 50% ‘light’ DNA 50% ‘medium’ DNA Matthew Meselson & Franklin Stahl 1958

DNA replication is semi-conservative!

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Conservative or semiconservative? Replication is semiconservative

DNA replication Replication is semi- conservative Helicase unwinds the double helix and breaks the hydrogen bonds DNA polymerase forms the new strands. Complimentary base pairing ensures that the new strands are the same as the original Two double helixes result, connected at the centromere

DNA replication DNA replicates prior to mitosis and meiosis. The sequence (order) of bases is preserved with each cell cycle. Every cell of your body contains the same genetic information.

1.Consider the process that a cell uses to replicate its double-stranded DNA to make copies for daughter cells. Which statement describes the DNA in daughter cells? A. The double helix in one daughter cell consists of two strands that were originally in the parent cell, while the double helix in the other daughter cell consists of two newly made strands. B. The two strands of the double helices in both daughter cells consist of segments of new and parental DNA. C. The double helices in each daughter cell consist of one parental strand and one newly made strand.

2.Which term describes DNA replication? A. Conservative B. Semiconservative C. Dispersive

3.What kinds of bonds link the two strands of a double helix to each other? A. Covalent bonds B. Hydrogen bonds C. Ionic bonds

4. How often does DNA polymerase make an error during DNA replication? A. One error in every 100 nucleotide pairs B. One error in every 1,000 nucleotide pairs C. One error in every 10,000,000 nucleotide pairs

5. Which enzyme adds nucleotides to form a new strand of DNA A. Helicase B. DNA polymerase C. RNA polymerase

C 1.Consider the process that a cell uses to replicate its double-stranded DNA to make copies for daughter cells. Which statement describes the DNA in daughter cells? A. The double helix in one daughter cell consists of two strands that were originally in the parent cell, while the double helix in the other daughter cell consists of two newly made strands. B. The two strands of the double helices in both daughter cells consist of segments of new and parental DNA. C. The double helices in each daughter cell consist of one parental strand and one newly made strand.

B 2.Which term describes DNA replication? A. Conservative B. Semiconservative C. Dispersive

B 3.What kinds of bonds link the two strands of a double helix to each other? A. Covalent bonds B. Hydrogen bonds C. Ionic bonds

C 4. How often does DNA polymerase make an error during DNA replication? A. One error in every 100 nucleotide pairs B. One error in every 1,000 nucleotide pairs C. One error in every 10,000,000 nucleotide pairs

B 5. Which enzyme adds nucleotides to form a new strand of DNA A. Helicase B. DNA polymerase C. RNA polymerase

How does DNA replicate? Each DNA strand acts as a template. Free nucleotides form new strands of DNA through complementary base pairing, adenine with thymine and cytosine with guanine. Through this process, the genetic sequence is maintained through generations of cells and generations of organisms. The process is more complex than this as replication can only occur in a 5’-3’ direction and enzymes are involved in catalysing this chemical process.

Suggest reasons why there is complimentary base pairing: C with G and A with T

DNA replication [Advanced] Nucleotides DNA Polymerase DNA Polymerase DNA replication is semi-conservative; Unwinding of double helix Hydrogen bonds between two strands are broken; Each strand of parent DNA used as template for synthesis; Synthesis continuous on leading strand but not continuous on lagging strand; Leading to formation of Okazaki fragments (on lagging strand); Synthesis occurs in 5´ → 3´ direction; RNA primer synthesized on parent DNA using RNA primase; DNA polymerase adds the nucleotides (to the 3´ end) added according to complementary base pairing; DNA polymerase removes the RNA primers and replaces them with DNA; DNA ligase joins Okazaki fragments; DNA replication [Advanced]