DNA Replication

proposed that chromosome (- a complex of DNA and protein) are composed of genes

test whether genes were made of protein or DNA by studying how a virus called T2 infects bacterium e. coli. Was it protein of DNA the genetic material that entered he bacterium?

proteins contain sulfur (but not phosphorus), and DNA contains phosphorus (but not sulfur)

1. Labeled viruses infect E. coli cultures. 2. Cultures are agitated in a kitchen blender to separate empty vial capsids from bacterial cells in culture. 3. Bacterial cells are spun down into pellets by centrifuation.

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a virus that attacks E. coli, consists almost entirely of a DNA core packed in a protein coat.

The Hershey-Chase experiment determined which part of the virus (protein or DNA) entered the bacterium.

1. labeled some viruses with radioactive sulfur, which attaches to proteins but not in DNA. 2. labeled with radioactive phosphorus, which attaches to DNA but not proteins.

1. The labeled phophorus (& thus the viral DNA) remained with the bacterial cells in pellet. 2. The labeled sulfur (& thus the viral protein) separated from the bacteria attached to capsid found in supernatant.

A protein must be the information molecule.

i. Semiconservative, Conservative replication, Dispersive replication

the parental DNA strands separate and each is used as a template for the synthesis of a new strand. Daughter molecules each consist of one old and one new strand.

The parental molecule serves as a template for the synthesis of an entirely new molecule.

the parent molecule is cut into sections such that the daughter molecules contain old DNA interspersed with newly synthesized DNA.

to provide more information about which DNA replication model was correct.

1. They grew E. coli in the presence of "heavy" nitrogen 15N (dense) to label the bacteria's DNA and collected an initial sample (Template DNA) 2. Next, they moved the bacteria to a normal 14N-containing medium (less dense) and collected samples from each bacterial generation. 3. DNA was extracted from each sample and separated by density gradient centrifugation.

The densities of the resulting DNA samples supported semiconservative DNA replication as the mechanism by which the hereditary material is duplicated.

DNA replicates dispersively.

DNA replicates conservatively.

No completely "heavy" DNA is observed after the first round of replication.

breaks the hydrogen bonds between the parental DNA strands and unwinds the double helix separating them

bind to the single strands of DNA, preventing them from re-forming hydrogen bonds with each other and allowing synthesis to occur on both strands.

cuts, swivels, and rejoins DNA strands ahead of the replication fork (downstream of the replication fork), relieving the tension farther down the helix that is caused by the unwinding of the DNA.

creates short RNA primers, initiating DNA synthesis on both template strands.

1. Helicase seperates/ unwinds DNA strands 2. SSBPs attach to seperated strands to prevent from closing 3. Topoisomerase relieves tension caused by replication fork 4. Primase synthesizes RNA primer 5. DNA polymerase III then adds bases to the 3' end of the primer continuously.

1. Helicase seperates/ unwinds DNA strands 2. SSBPs attach to seperated strands to prevent from closing 3. Topoisomerase relieves tension caused by replication fork 4. primase synthesizes a RNA primer. 5. polymerase III then adds bases to the 3' end of the primer. 6. The lagging strand is synthesized as short discontinuous fragments called Okazaki fragments. 7. DNA polymerase I removes the primers and fills the gaps 8. ligase seals Okazaki fragments

the enzyme that catalyzes DNA synthesis adding nucleotides to only the 3′ end of a growing DNA chain (from 5' to 3' direction)

Energy comes from breaking the bonds between the three phosphates of incoming dNTPs.

is primarily responsible for chromosomal DNA replication. Synthesizes the new strands, but it requires an existing 3′ hydroxyl (—OH) group to add nucleotides.

- Are involved in primer removal and DNA repair. - Removes the RNA primer at the beginning of each Okazaki fragment and fills in the gap. - removes the RNA primers and replaces them with DNA nucleotides.

forms in a chromosome that is actively being replicated. Grow as DNA replication proceeds.

is the Y-shaped region where the DNA is split into two separate strands for copying.

In bacterial chromosomes, is the location where the replication process begins

eukaryotes and prokaryotes have bidirectional replications: DNA replicates in both directions from the origin, forming two replication forks. In DNA replication, both strands of DNA act as templates.

It leads into the replication fork and is synthesized continuously in the 5′ → 3′ direction.

is synthesized discontinuously in the direction away from the replication fork and so lags behind the fork. Synthesis of the lagging strand away from the fork occurs because DNA synthesis must proceed in the 5' → 3' direction.

short discontinuous fragments made in the synthesis of the lagging strand. Synthesised independently and later joined together.

another name for lagging strand. Named because of the formation of Okazaki fragments

1. DNA polymerase I removes the RNA primer at the beginning of each Okazaki fragment and fills in the gap 2. DNA ligase then joins the Okazaki fragments to form a continuous DNA strand.

joins Okazaki fragments by forming phosphodiester bonds between them, thus completing DNA replication on lagging strand.

1. Primase adds RNA primer 2. DNA polymerase III extends the segment 3. DNA polymerase I removes the primer 4. ligase seals the gaps.

1. Helicase seperates/ unwinds DNA strands 2. SSBPs attach to seperated strands to prevent from closing 3. Topoisomerase relieves tension caused by replication fork 4. Primase synthesizes primer 5. DNA polymerase III adds bases and elongates strand from 5' to 3'.

multi-enzyme machine responsible for DNA synthesis around the replication fork

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the lagging strand

end of the chromosome after DNA replication generally consist of short non coding repeating stretches of bases

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base pairing cannot be completed at this regions, resulting in shorter strands

DNA polymerase can only add nucleotides at the 3' end (5' to 3' direction). So the extension of the lagging strand outward from the fork. At the end of the strand there will be no space for primer to bind and this will result in the lagging strand remaining shorter. With consecutive rounds of DNA replication, the ends of the DNA (telomeres, will become shorter and shorter.

telomerase (protein and RNA) can reverse transcribe onto its own RNA template to synthesize the additional DNA making the 3' telomere longer. DNA polymerase III and Okasaki fragments can now extend the 5' end of the other DNA strand. There will always be some 3' single stranded DNA at the end, but telomerase will ensure that the telomeres maintain their original length and that genes on the chromosome are protected.

Telomerase is involved in adding DNA to the end of the lagging strand. At the ends of linear chromosomes (telomeres), telomerase uses its built-in RNA template to extend the parent DNA template near the end of the lagging strand, providing room for an RNA primer so that lagging strand synthesis can be completed to the end of the chromosome

prevent the loss of DNA from chromosome ends.

Both DNA polymerase and primase are involved in synthesizing the leading and lagging strands.

the two ends of the strand have different chemical groups

¾ low-density and ¼ intermediate-density. Semiconservative replication of the second generation DNA would produce a ratio of 2 hybrid to 6 14N double helices (1:3).

in somatic cells there is no mechanism to prevent telomeres from becoming shorter after consecutive replications. Do not express tolomerase so over time the chromosmes in these cells will shorten (aging)

prevent lagging strand from getting shorter after every replication. Stem cells in bone marrow are rapidly dividing cells that need to express telomerase to prevent chromosome from becoming shorter as this could eventually impact the genes in the cell.

One of the changes found in tumor cells which are rapid dividing cells. If within rapidly dividing cells, the chromosomes become shorter and shorter it would ultimately effect some important gene required for replication and cell division would stop. Cancer cells tend to have mutations in the polymerase genes, expressing telomerase, allowing for more cell division.

. Human DNA contains more origins of replication than prokaryotic DNA.

DNA ligase