3. Experiments by Matthew Meselsohn and Franklin Stahl proved DNA replication was semi conservative. Using Esherichia coli (bacterium), they used two isotopes of nitrogen and observed which isotopes were present in the stands of DNA before and after replication.

4. http://highered.mheducation.com/sites/dl/free/00728 35125/126997/animation16.html http://highered.mheducation.com/sites/dl/free/00728 35125/126997/animation16.html TASK: read pages 112/3 and answer the DBQ

5. DNA replication always occurs just before mitosis. 4000 nucleotides can be replicated per second! Some bacteria divide every 20 minutes Far more nucleotides in eukaryotic cells compared to prokaryotic cells Very few errors (mutations) occur during DNA replication

6. Origin of Replication – site DNA replication begins. Prokaryotes have a single origin, Eukaryotes have thousands of origins. More origins you have, the faster DNA can replicate. 1. Enzyme helicase separates strands (breaks H bonds between bases) – bubble appears. 2. Replication fork at each end of bubble. DNA forms two parental DNA strands (templates) 3. Bubble extends in both directions (bidirectional). Parallel strands produced (Daughter strands)

7. http://highered.mheducation.com/sites/0072943696/ student_view0/chapter3/animation__dna_replication __quiz_1_.html http://highered.mheducation.com/sites/0072943696/ student_view0/chapter3/animation__dna_replication __quiz_1_.html

8. 1. Primase produces a primer (short sequence of RNA, 5-10 nucleotides). Primase allows RNA nucleotides to join to exposed DNA bases. 2. DNA polymerase III allows additional DNA nucleotides to join DNA strand. Strand grows in a 5’ to 3’ direction. 3. DNA polymerase I removes primer from 5’ end. Primer put with DNA nucleotides.

9. Remember… DNA can only be assembled in the 5’ to 3’ direction because this is the direction polymerase III works in. So, the two strands assemble differently The 3’ to 5’ template strand, the new DNA strand is formed as just described. Process is very fast and continuous. Known as The Leading Strand. The other strand forms more slowly, known as The Lagging Strand.

10. The reason the lagging strand forms more slowly is because it is assembled in fragments. Okazaki fragments. Enzyme DNA ligase joins fragments together, forming a continuous strand

11. You should be able to draw and annotate this diagram. Or the version in the text book. Page 200 http://highered.mcgraw- hill.com/sites/0072943696/student_view0/chapter3/animation__dna_replica tion__quiz_1_.html

12. ProteinRole HelicaseUnwinds double helix at replication fork PrimaseSynthesises RNA primer DNA polymerase IIISynthesises new strand by adding nucleotides onto primer. 5’ to 3’ direction DNA polymerase IRemoves primer, replaces with DNA DNA ligaseJoins end of DNA segments and Okazaki fragments Summary of proteins. *when helicase is ‘unzipping’ DNA at multiple site along a strand, the enzyme topoisomerase ensures the strand remains stable.

13. The STEPS 1.Helicase uncoils the DNA 2.RNA primase adds short sequences of RNA to both strands (the primer) 3.The primer allows DNA polymerase III to bind and start replication 4.DNA polymerase III adds nucleotides to each template strand in a 5' → 3' direction 5.These nucleotides are initially deoxyribonucleoside triphosphates but they lose two phosphate groups during the replication process to release energy 6.One strand is replicated in a continuous manner in the same direction as the replication fork (leading strand) 7.The other strand is replicated in fragments (Okazaki fragments) in the opposite direction (lagging strand) 8.DNA polymerase I removes the RNA primers and replaces them with DNA 9.DNA ligase then joins the Okazaki fragments together to form a continuous strand

14. Another summary DNA replication is semi-conservative and occurs during the S phase of interphase Helicase unwinds and separates the double stranded DNA by breaking the hydrogen bonds between base pairs This occurs at specific regions (replication origins), creating a replication fork of two polynucleotide strands in antiparallel directions RNA primase synthesises a short RNA primer on each template strand to provide an attachment and initiation point for DNA polymerase III DNA polymerase III adds deoxynucleoside triphosphates (dNTPs) to the 3' end of the polynucleotide chain, synthesising in a 5' - 3' direction The dNTPs pair up opposite their complementary base partner (adenine pairs with thymine ; guanine pairs with cytosine) As the dNTPs join with the DNA chain, two phosphates are broken off, releasing the energy needed to form a phosphodiester bond Synthesis is continuous on the strand moving towards the replication fork (leading strand) Synthesis is discontinuous on the strand moving away from the replication fork (lagging strand) leading to the formation of Okazaki fragments DNA polymerase I removes the RNA primers and replaces them with DNA DNA ligase joins the Okazaki fragments together to create a continuous strand

15. Some animations on the site below: http://highered.mcgraw- hill.com/sites/0072943696/student_view0/chapter3/animation__dna_replication__quiz_1_.html These might also be useful? http://www.wiley.com/college/pratt/0471393878/student/animations/dna_replication/inde x.html http://www.johnkyrk.com/DNAreplication.html http://bioteach.ubc.ca/TeachingResources/MolecularBiology/DNAReplication.swf http://learn.genetics.utah.edu/content/begin/dna/builddna/ http://www.stolaf.edu/people/giannini/flashanimat/molgenetics/dna-rna2.swf http://www.courses.fas.harvard.edu/~biotext/animations/replication1.swf Theory is on the next few slides