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”

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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

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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’

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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

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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….

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