Avery, McCarty, and MacLeod ExperimentWhat were their conclusion?

CarbohydratesCarbohydrates LipidLipidss

ProteinProteinss

RNARNA DNADNA

Mice were given deadly bacteria with Mice were given deadly bacteria with enzymes that destroyed …enzymes that destroyed …

S-Type S-Type Carbohydrates Carbohydrates

DestroyedDestroyed

S-Type S-Type Lipids Lipids

DestroyeDestroyedd

S-Type S-Type Proteins Proteins DestroyeDestroye

dd

S-Type S-Type RNA RNA

DestroyeDestroyedd

S-Type S-Type DNA DNA

DestroyeDestroyedd

Conclusion:Conclusion:DNA was the DNA was the

transforming factor! transforming factor! DNA DNA is the molecule of is the molecule of

inheritance inheritance

Learning OutcomesContent• Explain which experiment demonstrated that

DNA was the molecule of inheritance• Investigate the theories of replication

Process• Conclusion to experimentation

Benefit• Explain how experimentation can make leaps

forward in our understanding.

3 proposed theories of how DNA replicates....

Any ideas to test these?Take an educated guess of which model and WHY!

N15 sole sourceof nitrogen for

many generationsCollect and analyze

DNA

Cell Transfer

Transfer cells to N14 andAllow cells to divide only once

Collect and analyze DNA

Allow cells to divide a second time

Collect and analyze DNA

Prediction Prediction Prediction

ConservativeAll heave labeled

DNASemi conservative

Not Relevant

ConservativeFirst generation

½ labeled with onlyheavy label

½ labeled with N14

Semi conservativeAll DNA hybrids

N15/N14

ConservativeOnly one heavy DNA

3 DNA N14

Semi conservative½ DNA N15

½ DNA N14

Meselson – Stahl Experiment

• To answer this question Meselson and Stahl exploited the differences between N14 and N15

– N15 is a heavy isotope with one extra neutron– The difference in mass creates a difference in

density– This difference in density can be seen when

centrifugation techniques are employed

DNA (helicase) unwinds to form 2 templates

(RNA) primer is then bound to the starting DNA site.

Begin of replication, an enzyme called DNA polymerase binds to the (RNA) primer, which indicates the starting point for the replication. Occurs at several locations on DNA strand

DNA polymerase can only synthesize new DNA

The DNA polymerase can only travel on one side of the original strand without any interruption (3’ to 5’).

Since the DNA replication on the lagging strand is not continuous, the replicated DNA is fragmented with fragments added by the enzyme ligase.

Semi-conservative replication!

Activity – Artists Easel

• Now draw the parts for each part I have described.

Self Check

• Compare what I’ve drawn to your own diagram/pictures ....

DNA ReplicationFrom this discovery the mechanism of how DNA replicates was proposed ...

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• Begins as DNA unwinds Begins as DNA unwinds (helicase(helicase))

• Two strands open forming Two strands open forming Replication Forks (Y-shaped Replication Forks (Y-shaped region)region)

• New strands grow at the forksNew strands grow at the forksReplicationReplicationForkFork

Parental DNA MoleculeParental DNA Molecule

3’

5’

3’

5’

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DNA ReplicationDNA Replication• DNA polymeraseDNA polymerase can only add can only add

nucleotides to the nucleotides to the 3’ end3’ end of the of the DNA DNA

• This causes the This causes the NEWNEW strand to be strand to be built in a built in a 5’ to 3’ direction5’ to 3’ direction

RNARNAPrimerPrimerDNA PolymeraseDNA PolymeraseNucleotideNucleotide

5’

5’ 3’

Direction of ReplicationDirection of Replication

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DNA ReplicationDNA Replication• As the 2 DNA strands open at As the 2 DNA strands open at

the origin, the origin, Replication Replication BubblesBubbles form form

• Meaning DNA replication Meaning DNA replication occurs at several locations occurs at several locations

Bubbles Bubbles

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Synthesis of the New DNA Synthesis of the New DNA StrandsStrands

• The The Leading StrandLeading Strand is synthesized as a single strand single strand from the point of origin toward the opening replication fork

RNARNAPrimerPrimerDNA PolymeraseDNA PolymeraseNucleotidesNucleotides

3’5’

5’

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Synthesis of the New DNA Synthesis of the New DNA StrandsStrands

• The The Lagging StrandLagging Strand is is synthesized discontinuouslydiscontinuously against overall direction of replication

• This strand is made in MANY short segments It is replicated from the replication fork toward the origin

RNA PrimerRNA Primer

Leading StrandLeading Strand

DNA PolymeraseDNA Polymerase

5’

5’

3’

3’

Lagging StrandLagging Strand

5’

5’

3’

3’

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Lagging Strand SegmentsLagging Strand Segments• ((Okazaki FragmentsOkazaki Fragments)) - - series of

short segments on the lagging lagging strandstrand

• Must be joined together by an Must be joined together by an enzyme Ligaseenzyme Ligase

Lagging Strand

RNARNAPrimerPrimer

DNADNAPolymerasePolymerase

3’

3’

5’

5’

Okazaki FragmentOkazaki Fragment

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Joining of Okazaki FragmentsJoining of Okazaki Fragments• The enzyme The enzyme LigaseLigase joins the joins the

Okazaki fragments together to Okazaki fragments together to make one strandmake one strand

Lagging Strand

Okazaki Fragment 2Okazaki Fragment 2

DNA ligaseDNA ligaseOkazaki Fragment 1Okazaki Fragment 1

5’

5’

3’

3’

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Replication of StrandsReplication of StrandsReplication Fork

Point of Origin

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Question:Question:•What would be the

complementary DNA strand for the following DNA sequence?

DNA 5’-CGTATG-3’DNA 5’-CGTATG-3’copyright cmassengale

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Answer:Answer:

DNA 5’-CGTATG-3’DNA 5’-CGTATG-3’DNA 3’-GCATAC-5’DNA 3’-GCATAC-5’

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Activity

• Add a series of diagrams/cartoons which explain the terms

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Proofreading New DNAProofreading New DNA• DNA polymerase initially makes DNA polymerase initially makes

about about 1 in 10,0001 in 10,000 base pairing base pairing errorserrors

• EnzymesEnzymes proofread and correct proofread and correct these mistakesthese mistakes

• The new error rate for DNA that The new error rate for DNA that has been proofread is has been proofread is 1 in 1 billion1 in 1 billion base pairing errorsbase pairing errors

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Semiconservative Model of Semiconservative Model of ReplicationReplication• Idea presented by Idea presented by Watson & CrickWatson & Crick• TheThe two strands of the parental

molecule separate, and each acts as a template for a new complementary strand

• New DNA consists of 1 PARENTAL (original) and 1 NEW strand of DNA

Parental DNA

DNA Template

New DNA

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