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DNA and Replication
DNA: The Primary Source of Heritable Information
• Genetic information is transmitted from one generation to the next through DNA or RNA
Chromosomes
• Non-eukaryotic (bacteria) organisms have circular chromosomes
• Eukaryotic organisms have multiple linear chromosomes
• Exceptions:
– Some bacterial cells have linear chromosomes
– Mitochondria and chloroplasts have been found to have circular and linear chromosomes
Plasmids
• Prokaryotes, viruses and eukaryotes (yeast) may contain plasmids
• Plasmids are small extra-chromosomal, double-stranded circular DNA molecules
• Plasmids make excellent cloning vectors
Plasmids as Cloning Vectors
Important Historical Experiments
• The proof that DNA is the carrier of genetic information involved a number of important historical experiments, including:
– Frederick Griffith
–Avery-MacLeod-McCarty
–Hershey-Chase
–Watson, Crick, Wilkins and Franklin
Frederick Griffith • 1928 - Experiments in bacterial transformation
• Smooth (S) pathogenic bacteria
• Rough (R) nonpathogenic bacteria
Frederick Griffith • Living bacterial cells were converted to
disease causing bacteria (transformation)
• “Transforming factor”
ANIMATION
Avery-MacLeod-McCarty Experiments
• Experiments demonstrated that DNA is the “transforming” material, not protein
Avery-MacLeod-McCarty Experiments
• Used different enzymes to destroy protein, RNA or DNA in separate tubes to determine if transformation occurs
Hershey-Chase Experiment
• 1952 - Concluded that DNA, not protein, functions as the genetic material of phage T2
Hershey-Chase Experiment
• Tagged protein coat with radioactive sulfur
• Tagged viral genome with radioactive phosphorus
ANIMATION
Erwin Chargaff
• 1952 - Nitrogenous base composition
• % of adenine is equal to % of thymine
• % of guanine is equal to % cytosine
• Composition of DNA varies from species to species
Maurice Wilkins and Rosalind Franklin
• 1951 - Worked with a technique called X-ray diffraction
• Determined the helical nature of DNA
James Watson and Francis Crick
• 1953 – Determined the structure of DNA using Chargaff’s and Franklin’s data
• Franklin’s picture showed two strands of nucleotides
DNA (Deoxyribonucleic acid)
• Nucleic acid
• Consists of monomers called nucleotides
• Stores genetic information, determines an organisms traits by synthesizing proteins
• Each organisms genome is unique
Structure of DNA
• Double helix
• Consists of a double strand of nucleotides
• Two strands are anti-parallel: strands are oriented in opposite directions
–5’ to 3’
–3’ to 5’
ANIMATION
Nucleotide Composition
• Three parts of a nucleotide
– 5 carbon sugar called deoxyribose
–Phosphate group
–A single nitrogenous base
Four Nitrogenous Bases
• Purines (double ring)
– Adenine (A)
– Guanine (G)
• Pyrimidines (single ring)
– Thymine (T)
– Cytosine (C)
Purines pair with Pyrimidines
Chargaff’s Rules
• A-T (2 hydrogen bonds)
• C-G (3 hydrogen bonds)
DNA Replication
Why do cells need to replicate DNA?
Semi-Conservative Replication
• New DNA molecules have one original template strand and one new strand
• Follows complementary base pair rules and begins at sites called origins of replication
• “Leading strand” – continuously synthesized
• “Lagging strand” – synthesized in fragments
ANIMATION
Semi-conservative Replication
Origin of Replication - Prokaryotes
Origins of Replication - Eukaryotes
Replication Bubbles
Enzyme Functions
• Helicase
• RNA Primase
• DNA Polymerase III
• DNA Polymerase I
• DNA Ligase
• Topoisomerase
• Single-strand binding proteins (not an enzyme)
Replication Direction
• Replication proceeds in the 5’ to 3’ direction
• DNA polymerase can only add free nucleotides to the 3' end of the newly forming strand
DNA Replication Animation
Steps of DNA Replication
More animations for your viewing pleasure….
ANIMATION