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er 16: The Molecular Basis of Inheritance apter 14 – traits carried in gametes apter 15 – genes found on chromosomes apter 16 – DNA is the molecule of inheritance w did we learn this?

Chapter 16: The Molecular Basis of Inheritance

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Chapter 16: The Molecular Basis of Inheritance. Chapter 14 – traits carried in gametes Chapter 15 – genes found on chromosomes Chapter 16 – DNA is the molecule of inheritance How did we learn this?. - PowerPoint PPT Presentation

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Page 1: Chapter 16:  The Molecular Basis of Inheritance

Chapter 16: The Molecular Basis of Inheritance

Chapter 14 – traits carried in gametes

Chapter 15 – genes found on chromosomes

Chapter 16 – DNA is the molecule of inheritance

How did we learn this?

Page 2: Chapter 16:  The Molecular Basis of Inheritance

Chapter 16: The Molecular Basis of Inheritance1. What evidence did Frederick Griffith have that DNA was the genetic material?

Bacteria of the “S” (smooth) strain of Streptococcus pneumoniae are pathogenic because they have a capsule that protects them from an animal’s defense system. Bacteria of the “R” (rough) strain lack a capsule and are nonpathogenic. Frederick Griffith injected mice with the two strains as shown below:

Griffith concluded that the living R bacteria had been transformed into pathogenic S bacteria by an unknown, heritable substance from the dead S cells.

EXPERIMENT

RESULTS

CONCLUSION

Living S(control) cells

Living R(control) cells

Heat-killed(control) S cells

Mixture of heat-killed S cellsand living R cells

Mouse dies Mouse healthy Mouse healthy Mouse dies

Living S cellsare found inblood sample.

Transformation – external DNA assimilated by a cell which changes the cell’s genotype and phenotype

Page 3: Chapter 16:  The Molecular Basis of Inheritance

Chapter 16: The Molecular Basis of Inheritance1. What evidence did Frederick Griffith have that DNA was the genetic material?2. How did Hershey & Chase determine that DNA was the genetic material?

- Bacteriophage – viruses that infect bacteria

Phagehead

Tail

Tail fiber

DNA

Bacterialcell

100

nm

Page 4: Chapter 16:  The Molecular Basis of Inheritance

Chapter 16: The Molecular Basis of Inheritance1. What evidence did Frederick Griffith have that DNA was the genetic material?2. How did Hershey & Chase determine that DNA was the genetic material?

In their famous 1952 experiment, Alfred Hershey and Martha Chase used radioactive sulfur and phosphorus to trace the fates of the protein and DNA, respectively, of T2 phages that infected bacterial cells.

EXPERIMENT

Radioactivity(phage protein)in liquid

Phage

Bacterial cell

Radioactiveprotein

Emptyprotein shell

PhageDNA

DNA

Centrifuge

Pellet (bacterialcells and contents)

RadioactiveDNA

Centrifuge

PelletRadioactivity(phage DNA)in pellet

Batch 1: Phages weregrown with radioactivesulfur (35S), which wasincorporated into phageprotein (pink).

Batch 2: Phages weregrown with radioactivephosphorus (32P), which was incorporated into phage DNA (blue).

1 2 3 4Agitated in a blender toseparate phages outsidethe bacteria from thebacterial cells.

Mixed radioactivelylabeled phages withbacteria. The phagesinfected the bacterial cells.

Centrifuged the mixtureso that bacteria formeda pellet at the bottom ofthe test tube.

Measured theradioactivity inthe pellet and the liquid

Page 5: Chapter 16:  The Molecular Basis of Inheritance

Chapter 16: The Molecular Basis of Inheritance1. What evidence did Frederick Griffith have that DNA was the genetic material?2. How did Hershey & Chase determine that DNA was the genetic material?

Phage proteins remained outside the bacterial cells during infection, while phage DNA entered the cells. When cultured, bacterial cells with radioactive phage DNA released new phages with some radioactive phosphorus.

Hershey and Chase concluded that DNA, not protein, functions as the T2 phage’s genetic material.

RESULTS

CONCLUSION

Page 6: Chapter 16:  The Molecular Basis of Inheritance

Chapter 16: The Molecular Basis of Inheritance1. What evidence did Frederick Griffith have that DNA was the genetic material?2. How did Hershey & Chase determine that DNA was the genetic material?3. What was Chargaff’s contribution?

- Nucleotide composition- Human

- A – 30.3%- T – 30.3%- C – 19.9%- G – 19.5%

Chargaff’s rulesA=TC=G

Sugar-phosphatebackbone

Nitrogenousbases

5 endO–

O P O CH2

5

4O–

HH

OH

HH

3

1H O

CH3

N

O

NH

Thymine (T)

O

O P O

O–

CH2

HH

OH

HH

HN

N

N

H

NH

H

Adenine (A)O

O P O

O–

CH2

HH

OH

HH

HH H

HN

NN

OCytosine (C)

O

O P O CH2

5

4O–

H

O

HH

3

1

OH2

H

N

NN H

ON

N HH

H H

Sugar (deoxyribose)3 end

Phosphate

Guanine (G)

DNA nucleotide

2

N

Note: 5’ end – phosphate group3’ end - -OH

Page 7: Chapter 16:  The Molecular Basis of Inheritance

Chapter 16: The Molecular Basis of Inheritance1. What evidence did Frederick Griffith have that DNA was the genetic material?2. How did Hershey & Chase determine that DNA was the genetic material?3. What was Chargaff’s contribution?4. What experiment did Watson & Crick do?

- NONE – compiled everyone else’s data

(a) Rosalind Franklin Franklin’s X-ray diffractionPhotograph of DNA

(b)Double helix

Page 8: Chapter 16:  The Molecular Basis of Inheritance

Chapter 16: The Molecular Basis of Inheritance1. What evidence did Frederick Griffith have that DNA was the genetic material?2. How did Hershey & Chase determine that DNA was the genetic material?3. What was Chargaff’s contribution?4. What experiment did Watson & Crick do?

- NONE – compiled everyone else’s data- Determined double helix from Rosalind Franklin’s data

Purine + Purine: too wide

Pyrimidine + pyrimidine: too narrow

Purine + pyrimidine: widthConsistent with X-ray data

Page 9: Chapter 16:  The Molecular Basis of Inheritance

Chapter 16: The Molecular Basis of Inheritance1. What evidence did Frederick Griffith have that DNA was the genetic material?2. How did Hershey & Chase determine that DNA was the genetic material?3. What was Chargaff’s contribution?4. What experiment did Watson & Crick do?

- NONE – compiled everyone else’s data- Determined double helix from Rosalind Franklin’s data

N H O CH3

N

N

O

N

N

N

N H

Sugar

Sugar

Adenine (A) Thymine (T)

N

N

N

NSugar

O H N

H

NH

N OHH

N

Sugar

Guanine (G) Cytosine (C)

Page 10: Chapter 16:  The Molecular Basis of Inheritance

O

–O O

OHP

H2C

O

–O O

OP

H2C

O

–O O

OP

H2C

O

–OO

OP

O

OH

O

O

OT A

C

GC

A T

O

O

O

O

OH

CH2

O O–

OOP

CH2

OO–

OOP

CH2

OO–

OO

P

CH2

OO–

OOP

5 end

Hydrogen bond3 end

5 end

3 end

C

T

A

A

T

CG

GC

A T

C G

AT

AT

A T

TA

C

TA0.34 nm

3.4 nm

(a) Key features of DNA structure (b) Partial chemical structure (c) Space-filling model

G

1 nm

G

H2C

G

Figure 16.7 The double helix

DNA is antiparallelHydrogen bonds hold double helix togetherNow that we know how DNA is shaped & how it fits together…..

Page 11: Chapter 16:  The Molecular Basis of Inheritance

Announcements- Test corrections

- Stapled behind- Place in box

- Canned food – no later than Thursday- Pie Festival – for Ch 16 – 18 test (instead of cans)

Page 12: Chapter 16:  The Molecular Basis of Inheritance

Chapter 16: The Molecular Basis of Inheritance1. What evidence did Frederick Griffith have that DNA was the genetic material?2. How did Hershey & Chase determine that DNA was the genetic material?3. What was Chargaff’s contribution?4. What experiment did Watson & Crick do? 5. How does DNA replicate itself? The basic concept

(a) The parent molecule has two complementary strands of DNA. Each base is paired by hydrogen bonding with its specific partner, A with T and G with C.

(b) The first step in replication is separation of the two DNA strands.

(c) Each parental strand now serves as a template that determines the order of nucleotides along a new, complementary strand.

(d) The nucleotides are connected to form the sugar-phosphate backbones of the new strands. Each “daughter” DNA molecule consists of one parental strand and one new strand.

ACTAG

ACTAG

ACTAG

ACTAG

TGATC

TGATC

ACTAG

AC

T

A

G

TGATC

TGATC

TGATC

T

G

A

TC

But is replication done in a conservative, semi-conservative or dispersive manner?

Page 13: Chapter 16:  The Molecular Basis of Inheritance

Conservativemodel. The twoparental strandsreassociate after acting astemplates fornew strands,thus restoringthe parentaldouble helix.

(a)

Semiconserva-tive model.The two strandsof the parentalmolecule separate, and each functionsas a templatefor synthesis ofa new, comple-mentary strand.

(b)

Dispersivemodel. Eachstrand of bothdaughter mol-ecules containsa mixture ofold and newlysynthesizedDNA.

(c)

Parent cellFirstreplication

Secondreplication

Figure 16.10 Three alternative models of DNA replication

Conservative- Original strands return together

Semi-conservative- One original & one new strand

Dispersive- Original strands are dispersed through new strands

The Meselson – Stahl Experiment showed how DNA replicates.

Page 14: Chapter 16:  The Molecular Basis of Inheritance

Matthew Meselson and Franklin Stahl cultured E. coli bacteria for several generations on a medium containing nucleotide precursors labeled with a heavy isotope of nitrogen, 15N. The bacteria incorporated the heavy nitrogen into their DNA. The scientists then transferred the bacteria to a medium with only 14N, the lighter, more common isotope of nitrogen. Any new DNA that the bacteria synthesized would be lighter than the parental DNA made in the 15N medium. Meselson and Stahl could distinguish DNA of different densities by centrifuging DNA extracted from the bacteria.

EXPERIMENT

The bands in these two centrifuge tubes represent the results of centrifuging two DNA samples from the flask in step 2, one sample taken after 20 minutes and one after 40 minutes.

RESULTS

Bacteriacultured inmediumcontaining15N

Bacteriatransferred tomediumcontaining14N

21

DNA samplecentrifugedafter 20 min(after firstreplication)

3 DNA samplecentrifugedafter 40 min(after secondreplication)

4Lessdense

Moredense

Figure 16.11 Does DNA replication follow the conservative, semiconservative, or dispersive model?

Page 15: Chapter 16:  The Molecular Basis of Inheritance

CONCLUSION Meselson and Stahl concluded that DNA replication follows the semiconservative model by comparing their result to the results predicted by each of the three models in Figure 16.10. The first replication in the 14N medium produced a band of hybrid (15N–14N) DNA. This result eliminated the conservative model. A second replication produced both light and hybrid DNA, a result that eliminated the dispersive model and supported the semiconservative model.

First replication Second replication

Conservativemodel

Semiconservativemodel

Dispersivemodel

Page 16: Chapter 16:  The Molecular Basis of Inheritance

Chapter 16: The Molecular Basis of Inheritance1. What evidence did Frederick Griffith have that DNA was the genetic material?2. How did Hershey & Chase determine that DNA was the genetic material?3. What was Chargaff’s contribution?4. What experiment did Watson & Crick do? 5. How does DNA replicate itself? 6. How is DNA replicated?

- Origin of replication - sequence of DNA

Page 17: Chapter 16:  The Molecular Basis of Inheritance

Chapter 16: The Molecular Basis of Inheritance1. What evidence did Frederick Griffith have that DNA was the genetic material?2. How did Hershey & Chase determine that DNA was the genetic material?3. What was Chargaff’s contribution?4. What experiment did Watson & Crick do? 5. How does DNA replicate itself? 6. How is DNA replicated?

- Origin- Elongation

- DNA polymerase- 500 bp/sec bacteria, 50 bp/sec in us- New base added to 3’end (-OH)- dNTPs provide energy

Page 18: Chapter 16:  The Molecular Basis of Inheritance

New strand Template strand5’ end 3’ end

Sugar A TBase

C

G

G

C

A

C

T

PP

P

OH

P P

5’ end 3’ end

5’ end 5’ end

A T

C

G

G

C

A

C

T

3’ end

Nucleosidetriphosphate

Pyrophosphate

2 P

OH

Phosphate

Figure 16.13 Incorporation of a nucleotide into a DNA strand

DNA polymeraseOH

Page 19: Chapter 16:  The Molecular Basis of Inheritance

Chapter 16: The Molecular Basis of Inheritance1. What evidence did Frederick Griffith have that DNA was the genetic material?2. How did Hershey & Chase determine that DNA was the genetic material?3. What was Chargaff’s contribution?4. What experiment did Watson & Crick do? 5. How does DNA replicate itself? 6. How is DNA replicated?

- Origin- Elongation- Dealing with antiparallel strands

Page 20: Chapter 16:  The Molecular Basis of Inheritance

Parental DNA

DNA pol Ill elongatesDNA strands only in the5 3 direction. 3

5

5

3

35

21

Okazakifragments

DNA pol III

Templatestrand

Leading strandLagging strand

32 1

Templatestrand DNA ligase

Overall direction of replication

One new strand, the leading strand,can elongate continuously 5 3as the replication fork progresses.

The other new strand, thelagging strand must grow in an overall3 5 direction by addition of shortsegments, Okazaki fragments, that grow5 3 (numbered here in the orderthey were made).

DNA ligase joins Okazakifragments by forming a bond betweentheir free ends. This results in a continuous strand.

2

3

1

4

Figure 16.14 Synthesis of leading and lagging strands during DNA replication

Page 21: Chapter 16:  The Molecular Basis of Inheritance

Overall direction of replication

3

3

3

35

35

35

35

35

35

35

3 5

5

1

1

21

12

5

5

12

35

DNA ligase forms a bond between the newest DNAand the adjacent DNA of fragment 1.

6 The lagging strand in this region is nowcomplete.

7

DNA pol 1 replaces the RNA with DNA, adding to the 3 end of fragment 2.

5

After the second fragment is primed. DNA pol III adds DNAnucleotides until it reaches the first primer and falls off.

4

After reaching the next RNA primer (not shown), DNA pol III falls off.

3

DNA pol III adds DNA nucleotides to the primer, forming an Okazaki fragment.

2

Primase joins RNA nucleotides into a primer.

1

Templatestrand

RNA primer

Okazakifragment

Figure 16.15 Synthesis of the lagging strand

Page 22: Chapter 16:  The Molecular Basis of Inheritance

Announcements- Learning log – available – greater percentage of “critical thinking” questions- People are hungry!!!!! – tardies????

Page 23: Chapter 16:  The Molecular Basis of Inheritance

Figure 16.16 A summary of bacterial DNA replicationOverall direction of replication

Helicase unwinds theparental double helix.

Molecules of single-strand binding proteinstabilize the unwoundtemplate strands.

The leading strand issynthesized continuously in the5 3 direction by DNA pol III.

Leadingstrand Origin of replication

Laggingstrand

Laggingstrand

LeadingstrandOVERVIEW

Leadingstrand

Replication fork

DNA pol III

Primase

PrimerDNA pol III Lagging

strand

DNA pol I DNA ligase

1

2 3

Primase begins synthesisof RNA primer for fifthOkazaki fragment.

4

DNA pol III is completing synthesis ofthe fourth fragment, when it reaches theRNA primer on the third fragment, it willdissociate, move to the replication fork,and add DNA nucleotides to the 3 endof the fifth fragment primer.

5 DNA pol I removes the primer from the 5 endof the second fragment, replacing it with DNAnucleotides that it adds one by one to the 3’ endof the third fragment. The replacement of thelast RNA nucleotide with DNA leaves the sugar-phosphate backbone with a free 3 end.

6 DNA ligase bondsthe 3 end of thesecond fragment tothe 5 end of the firstfragment.

7

Parental DNA

5

3

43

21

5

3

Page 24: Chapter 16:  The Molecular Basis of Inheritance

Chapter 16: The Molecular Basis of Inheritance1. What evidence did Frederick Griffith have that DNA was the genetic material?2. How did Hershey & Chase determine that DNA was the genetic material?3. What was Chargaff’s contribution?4. What experiment did Watson & Crick do? 5. How does DNA replicate itself? 6. How is DNA replicated?7. How is DNA repaired?

- Proofreading – mismatch repair – DNA polymerase fixes mistakes made during copying

- Excision repair

Page 25: Chapter 16:  The Molecular Basis of Inheritance

Nuclease

DNApolymerase

DNAligase

A thymine dimerdistorts the DNA molecule.1

Repair synthesis bya DNA polymerasefills in the missingnucleotides.

3

DNA ligase seals theFree end of the new DNATo the old DNA, making thestrand complete.

4

A nuclease enzyme cutsthe damaged DNA strandat two points and thedamaged section isremoved.

2

Figure 16.17 Nucleotide excision repair of DNA damage

Page 26: Chapter 16:  The Molecular Basis of Inheritance

Chapter 16: The Molecular Basis of Inheritance1. What evidence did Frederick Griffith have that DNA was the genetic material?2. How did Hershey & Chase determine that DNA was the genetic material?3. What was Chargaff’s contribution?4. What experiment did Watson & Crick do? 5. How does DNA replicate itself? 6. How is DNA replicated?7. How is DNA repaired?8. What happens at the end of chromosomes (telomere)?

- Recall DNA polymerase works 5’ → 3’ direction- After RNA primer is removed, chromosome is shortened

Page 27: Chapter 16:  The Molecular Basis of Inheritance

End of parentalDNA strands

Leading strandLagging strand

Last fragment Previous fragment

RNA primer

Lagging strand

Removal of primers andreplacement with DNAwhere a 3 end is available

Primer removed butcannot be replacedwith DNA because

no 3 end availablefor DNA polymerase

Second roundof replication

New leading strandNew lagging strand 5

Further roundsof replication

Shorter and shorterdaughter molecules

5

3

5

3

5

3

5

3

3

Figure 16.18 Shortening of the ends of linear DNA molecules

Page 28: Chapter 16:  The Molecular Basis of Inheritance

2009 Nobel Prize in Medicine - for the discovery of how chromosomes are protected by telomeres and the enzyme telomerase

Telomerase - active in germ line cells & newborns

to prevent shortening-extends telomere a bit-has its own RNA

- complementary to telomere- serves as primer