DNA Replication. Back ground.. Single celled zygote goes from being single celled to 100 trillion more cells in over 240 days in humans! Wow!

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1 DNA Replication Back ground.. Single celled zygote goes from being single celled to 100 trillion more cells in over 240 days in humans! Wow! Must be fast! six billion base pairs in a single human cell all replicated in a few hours. (single cell has about of DNA) Must make no mistake! There is about one error in a billion nucleotide base pairs. So lets find out how! 1

2 o Watson and Crick know that the DNA being complementary to each other was a means for replication. As DNA unwinds each strand acts as a template to build a new helix. o Other scientists.. 3 main possibilities of DNA replication 1. Conservative Theory o Two new daughter strands formed from parents strand and then the two new strands join together and the parents are thus. conserved 2

3 2. semi conservative theory o Daughter DNA is made up of one strand of the parent and one strand of the new DNA 3. dispersive theory o DNA parental is broken into smaller fragments and daughters are an assortment of parental and new DNA 3

4 Matthew Meselson Franklin Stahl

5 o Grew bacteria on a medium containing. 15N This was a nitrogen with a higher molecular mass of 15 (vs 14) o As the bacteria grew/developed they incorporated the heavier nitrogen o They Isolated and centrifuged the DNA Lab Results: Heavy Bacteria Explanation: As the bacteria developed, their DNA incorporated the heavy nitrogen making a lower band Light Bacteria 5

6 o The heavy bacteria are transferred to a medium of light/regular/14 nitrogen. After the colony doubles (one complete round of cell ) division Lab Results 1st Generation (one hybrid band) Explanation Ruled out Conservative theory because would have seen a heavy band and a light band 2nd Generation: (one regular band and one hybrid) ruled out dispersive because would have continue to see one hybrid band 6

7 DNA Replication DNA replication in bacteria is relatively simple due to circular shape 7

8 Process of Replication I. Initiation Phase II. Elongation III.Termination I. Initiation Phase Replication origin o sequences of base pairs in prokaryotes and a sequence of in eukaryotic cells o Recognized by a group of enzymes that bind to DNA o helicase separates the two strands creating a bubble 8

9 Replication Forks o Where parent strand separates Helicases o Enzymes cleave and unravel short segments of DNA just ahead of the replication fork SSB s (single strand binding proteins) o Bind to the single strands preventing reannealing o Primer is a short strand of RNA the serves as the starting point for attachment of new nucleotides. The primer is catalyzed by the enzyme primase. 9

10 DNA Polymerase o Inserts into the bubble o Adds nucleotides complementary to build new strands using parent strand as a template o Can only attach nucleotides to an existing nucleotide chain II. Elongation o DNA polymerase only attaches to the 3 end so the elongation or the copy occurs from 5' to 3' Question? But if DNA is formed from 5 to 3 how can both strands be replicated simultaneously? 10

11 o A second DNA polymerase replaces the RNA primer starting at 5 to 3 o The leading strand only has one primer but the lagging strand has a new primer for every Okazaki fragment. o Lagging strand must be spliced together by DNA ligase 11

12 III.Termination o Daughter DNA rewind automatically back to helical shape o At every 5' end once the RNA primer is removed, there is no adjacent nucleotide chain with a 3' end that can be extended to fill the gap 12

13 o The cell only has enzymes that work 3' to 5' on parental strand o unpaired complementary strand will break off with out the stability double helix shape. o Therefore every generation a small portion is lost. o Humans lose 100 base pairs every replication o Prokaryotes have circular DNA so no problem! o telomeres at ends of chromosomes are areas of highly repetitive DNA (nucleotide) portions..expect to provide a buffer 13

14 Finally.. Proof Reading is done by DNA polymerase as well. It checks for hydrogen bonding to ensure the placement of correct base pairs. 14

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