dna, the genetic material the story of the discovery of the structure and function of dna. this...

DNA, the Genetic Material

The story of the discovery of the Structure and Function of DNA.

This correlates with chapter 16 in your textbook

Big Picture

• For most of the last century, no one knew what Mendel’s “factors” were.– Not Mendel– Not Darwin– The major principals of Genetics and Evolution were worked out, but the “factors” remained unknown.

• Sutton first developed chromosomal theory in 1902, but there was only circumstantial evidence that factors were genes on chromosomes.

This is a story...

• About the process of science, as well as the structure and function of DNA.

• For many years scientists thought that ___ was the heritable material, but of course we now know that it is DNA.

What is the genetic material?The Hammerling Experiment: Cells Store Hereditary Information in the NucleusJoachim Hammerling discovered that hereditary information in a green alga resided in the foot region, which is also the location of the nucleus.

How did he know?

• By amputating different areas, grafting to other alga, and seeing what parts would grow.

• The form that was found in the foot region is the one that would influence new growth.

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Nucleus in base determinestype of cap regenerated

A. crenulata A. mediterranea

• Transplantation Experiments: Each Cell Contains a Full Set of Genetic Instructions Later experiments in the mid-1950s showed that the nucleus of eukaryotic cells includes a full set of genetic information. – What happens if you transplant the nucleus

of Frog A to the egg of Frog B? Which frog will develop?

• Totipotent- nucleus of adult cell carries instructions to create entire organism

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Egg

Tadpole orfrog embryo

UV light destroysnucleus, or it is removedwith micropipette.

Differentiated cells areisolated from tadpoleor frog embryo.

Nucleus isremovedin micropipette.

Differentiated cellnucleus is insertedinto enucleate egg.

No growth

Embryo

Embryo

Tadpole

Abnormalembryo

Occasionally,an adultfrog develops.

12

3

A

B

• The Griffith Experiment: Hereditary Information Can Pass Between Organisms

• 1928 Frederick Griffith -

• Non-pathogenic S. pneumoniae was transformed by dead pathogenic S. pneumoniae. Information specifiying the virulent surface protein had been delivered to the safe live form.

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Mice die; their bloodcontains live pathogenicstrain of S. pneumoniae

Mixture of heat-killedpathogenic and livenonpathogenic strainsof S. pneumoniae

+

Heat-killed pathogenicstrain of S. pneumoniae

Live pathogenicstrain of S. pneumoniae

Live nonpathogenicstrain of S. pneumoniae

Polysaccharidecoat

Mice liveMice die(1) (3) (4)

Mice live(2)

Transformation

• Griffith didn’t know the factor that was transmitted was DNA. He hypothesized it was protein.

• This is called transformation. We know DNA from the deadly strain was taken up by the non-deadly strain.

• We will transform E. coli with a glowing gene, and the bacteria should glow.

• The Avery and Hershey-Chase Experiments: The Active Principle Is DNA

• 1944- Oswald Avery, Colin MacLeod and Maclyn McCarty provided conclusive evidence that DNA is the hereditary material for the transformation under investigation.

– Repeated experiments similar to Griffith’s, but removed as much protein and other material as possible, but still non- pathogenic cells were transformed, so protein, fat, nor carb responsible.

Alfred Hershey and Martha Chase -

• Bacteriophage: virus with a simple nucleic acid and a protein coat.

• Identified DNA by marking phosphorous with 32P (only DNA contains phosphorous)

• Identified protein by marking sulfur with 35S (only protein contains sulfer).

• Found that it was the DNA, marked with 32P, that was injected into the bacteria that caused disease.

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Protein coat labeledwith 35S

DNA labeled with 32P

Bacteriophages infectbacterial cells.

T2 bacteriophagesare labeled withradioactive isotopes.

Bacterial cells are agitatedto remove protein coats.

35S radioactivityfound in the medium

32P radioactivity foundin the bacterial cells

• What is the structure of DNA?The Chemical Nature of Nucleic Acids Both DNA and RNA are formed of nucleotides joined together in series. Each nucleotide is composed of a five-carbon sugar, a phosphate group, and a nitrogen-containing base.

Chargaff's Rule - there are always equal proportions of purines and pyrimidines.

• A-T• G-C • Pure AGgie (purines are A & G)

– Purines have a double ring structure.

– Pyrimidines have a single ring structure. Single always pairs with double.

.

-Do you remember how we numbered carbons in sugar? Start from right side.

-Base attached to 1’ phosphate attached to 5’. This comes up when we talk about replication, transcription, and translation. (We read from 5’ to 3’)

Phosphodiester bond between nucleotides.

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OH

CH2

O

4

5

3 2

1

PO4

Base

Figure 16.7

3.4 nm

1 nm

0.34 nm

Hydrogen bond

(a) Key features ofDNA structure

Space-fillingmodel

(c)(b) Partial chemical structure

3 end

5 end

3 end

5 end

T

T

A

A

G

G

C

C

C

C

C

C

C

C

C

C

C

G

G

G

G

G

G

G

G

G

T

T

T

T

T

T

A

A

A

A

A

A

• The Three-Dimensional Structure of DNA (page 309) Rosalyn Franklin was able to obtain the first glimpse of DNA using X-ray diffraction in 1953, while Watson and Crick theorized that DNA exists in a double-helical, antiparallel configuration.

• Famous example of woman being scr’d.

• Could you guess the pattern by looking at the x-ray?

How does DNA replicate?The Meselson-Stahl Experiment: DNA Replication Is Semiconservative Matthew Meselson and Franklin Stahl demonstrated that DNA replication is semiconservative because each strand of the original duplex becomes one of the two strands in each new duplex. (p. 312)

• Also used isotopes, this time 15N, a heavy isotope of Nitrogen

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4. The DNA was suspendedin a cesium chloridesolution.

Centrifugation

1 2 3 4

Control group(unlabeled DNA)

Labeled parentDNA (both strandsheavy)

F1 generation DNA(one heavy/light hybridmolecule)

F2 generation DNA(one unlabeled molecule,one heavy/light hybridmolecule)

• What would the F1 generation look like if completely new DNA were synthesized (conservative model), rather than semi-conservative replication?

• The new piece would be completely light, and the old piece would be completely heavy. There would be two separate bands of DNA.

• The Replication Process (PAGE 315-317) Replication of E. coli begins at a specific origin, proceeds bidirectionally, and ends at a specific terminus.

• OriC is beginning point.

• Contains many A-T pairs, which are double bonded, easy to open.

• Leading and lagging strand, why so named?

• One side is synthesized continuously, but the other limited to short segments (Okizaki fragments).

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DNApolymerase III

DNA double helix

Lagging strand

Primer

Okazakifragment

Leadingstrand

59

39

Fig

ure

16.

15

Pag

e 31

5Leadingstrand

Laggingstrand

Overview

Origin of replication Laggingstrand

Leadingstrand

Primer

Overall directionsof replication

Origin of replication

RNA primer

Sliding clamp

DNA pol IIIParental DNA

35

5

33

5

3

5

3

5

3

5

• DNA primase -creates a short RNA primer complementary to a DNA template

• DNA helicase unwinds the helix

• DNA polymerase, which then synthesizes new DNA by adding nucleotides to the growing strands

• DNA ligase creates phosphodiester bonds between adjacent Okazaki fragments

• Each of these has a name that gives away its job.

• Replication fork- Open area of DNA where replication takes place.

• Topoisomerase- Relieves tension in the area ahead of replication fork

• Replication can be divided into three stages: initiation, elongation, and termination. – Initiation- there are two OriC’s, one on

each strand.

Figure 16.17PAG PAGE 317 Overview

Leadingstrand

Origin of replication Lagging

strand

LeadingstrandLagging

strand Overall directionsof replicationLeading strand

DNA pol III

DNA pol III Lagging strand

DNA pol I DNA ligase

PrimerPrimase

ParentalDNA

5

5

5

5

5

33

3

333 2 1

4

Page 317

DNA REPLICATION

• DNA Replication is directional.

• Synthesis of DNA and RNA occurs in the 5’ to 3’ direction.

• We are studying replication. What part of the cell cycle is replication?

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OH

CH2

O

• Eukaryotic DNA Replication The major difference between prokaryotic and eukaryotic replication is that eukaryotic chromosomes have multiple replication origins, whereas prokaryotic chromosomes have a single point of origin.

•What is a gene?The One-Gene/One-Polypeptide Hypothesis Beadle and Tatum concluded that genes produce their effects by specifying the structure of enzymes, and that each gene encodes the structure of one enzyme. Today, this is commonly referred to as the one-gene/one-polypeptide relationship.

Nutritional Mutants in Neurospora: Scientific Inquiry

• George Beadle and Edward Tatum exposed bread mold to X-rays, creating mutants that were unable to survive on minimal media

• Using crosses, they and their coworkers identified three classes of arginine-deficient mutants, each lacking a different enzyme necessary for synthesizing arginine

• They developed a one gene–one enzyme hypothesis, which states that each gene dictates production of a specific enzyme

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Figure 17.2

Minimal medium

No growth:Mutant cellscannot growand divide

Growth:Wild-typecells growingand dividing

EXPERIMENT RESULTS

CONCLUSION

Classes of Neurospora crassa

Wild type Class I mutants Class II mutants Class III mutants

Minimalmedium(MM)(control)

MM ornithine

MM citrulline

Con

diti

on

MM arginine(control)

Summaryof results

Can grow withor without anysupplements

Can grow onornithine,citrulline, orarginine

Can grow onlyon citrulline orarginine

Require arginineto grow

Wild type

Class I mutants(mutation in

gene A)

Class II mutants(mutation in

gene B)

Class III mutants(mutation in

gene C)

Gene (codes forenzyme)

Gene A

Gene B

Gene C

Precursor Precursor Precursor PrecursorEnzyme A Enzyme A Enzyme A Enzyme A

Enzyme B Enzyme B Enzyme B Enzyme B

Enzyme C Enzyme C Enzyme C Enzyme C

Ornithine Ornithine Ornithine Ornithine

Citrulline Citrulline Citrulline Citrulline

Arginine Arginine Arginine Arginine

Figure 17.2c

CONCLUSION

Wild type

Class I mutants(mutation in

gene A)

Class II mutants(mutation in

gene B)

Class III mutants(mutation in

gene C)

Gene (codes forenzyme)

Gene A

Gene B

Gene C

Precursor Precursor Precursor PrecursorEnzyme A Enzyme A Enzyme A Enzyme A

Enzyme B Enzyme B Enzyme B Enzyme B

Ornithine Ornithine Ornithine Ornithine

Enzyme C Enzyme C Enzyme CEnzyme C

Citrulline Citrulline Citrulline Citrulline

Arginine Arginine Arginine Arginine

• How DNA Encodes Protein Structure

• Sanger sequenced the amino acids of insulin, first time to sequence a protein. Then Ingram found that a single aa substitution Valine instead of Glutamic acid, caused sickle cell anemia.

Over 50 years of research has yielded clear evidence that DNA is the molecule

responsible for the inheritance of traits from one generation to the next, and that DNA is divided into functional subunits, or genes,

located on chromosomes

The question is answered