Download DNA is the Genetic Material

Lecture 1; 2007
Lecture#1
Section B2; Good
DNA is the Genetic Material
Readings:
Griffiths et al (2004) 8th Edition: Chap. 1, 2-4; Chap. 7 pp 227-249
Problems: Chap. 7: 1-25, 26, 27
Genetics has been approached from two directions. Mendel, Morgan and Sturtevant
were interested in genes in terms of how they encode traits and specifically, how they
are inherited. However, the other question was what are gene made of, from a chemical
perspective. Whatever the substance was, it must be capable of containing the
information necessary to convert one phenotype to another, and have that change be
permanently heritable.
What were the key experiments that lead us to the conclusion that DNA is the
Genetic Material?
1. Griffiths -1928 - Heat killed cells can transform bacteria. Genes are made of DNA?
2. Avery, MacLeod, & McCarty - 1944 - DNA transforms bacteria
3. Hershey & Chase - 1952 - DNA (not protein) is the genetic material
4. The structure of DNA is a double helix, which is compatible with its role as the
genetic material.
Frederick Griffith in 1928.
"Basic Research" with bacterium
Streptococcus pneumonia (diplococcus), which causes pneumonia in humans
2 strains: smooth (S) and rough (R).
Figure 7-2 (8th) 8-1(7th)
a) S - smooth colonies - virulent (causes pneumonia)
->inject -> kills mice
b) R - rough colonies - non-virulent (don't cause
pneumonia) ->inject -> live mouse
c) heat killed S-strain cells ->injected into-> mouse -> lives \ polysaccharide coat not
cause of death
d) -> heat killed S-strain cells + R-strain live cells -> dead mouse. (Whereas 30 control
mice R-strain lived)
Further Griffith"s found that:
- live cells could be recovered from these dead mice -> had - S-smooth colonies
- virulent upon subsequent injections
Conclude:
something in the S-cell debris could convert (transform) R -> S
-Process called transformation
- "Transforming Principle"
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Lecture 1; 2007
Section B2; Good
Figure 7-2. The presence of heat-killed S cells transforms live R cells into live S cells.
Notes: First, they did not need mice as the R and S phenotype was enough. Second,
you will do the same in the lab (transformation), but using a plasmid containing an
antibiotic resistance marker.
Avery, MacLeod & McCarty – 1944 Fig 7-3 (8th) 8-2(7th)
Purified cell molecules and test each for ability to transform
- purified the transforming substance
- chemical composition agreed with that of DNA
- had High MW (500,000)
- protease treatment -> activity intact
- treatment with ribonuclease -> activity intact
- treatment with deoxyribonuclease -> activity lost
Transforming substance is DNA -> DNA is the gene, add to R-cells -> S-cells DNA in
vitro - transform
But at the time there was reluctance to accept DNA as the genetic material
Transforming substance
- contaminated with other chemical that is really the activity
- proteins were thought to be more likely since the complexity of DNA is only 4 simple
bases. Proteins are more complex.
Class Question:
Q.) DNA may only induce a change - just a mutagenic agent?
Is it possible that DNA is not actually the genetic material itself?
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Lecture 1; 2007
Section B2; Good
Fig 7-3 (8th) 8-3(7th) DNA is the agent transforming the R strain into virulence.
Experiment of Hershey & Chase - 1952
Bacteriophage (bacterial virus) T2
- very simple -> Protein coat outside - DNA inside
Therefore the genetic material is either protein or DNA
Hershey & Chase used different radioisotopes to follow each of these components
during the process of infection of the bacteria and replication of the Phage.
Radioisotopes: 32P to follow DNA; P not found in protein 35S labels protein; S not found in
DNA Incorporated radioisotopes into phage in separate cultures
- infected E.coli with many virus per cell
- after sufficient time ->separated phage ghosts & cells
Result
S protein -> ghosts - protein never entered the cell 32P DNA -> did enter the cells
Furthermore 32P could be recovered from the phage progeny
---> For bacteriophage T2 the genetic information is in DNA not protein
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Conclusion:
1- DNA is the hereditary material
2- But this was still not immediately accepted as proof that genes were DNA.
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Lecture 1; 2007
Section B2; Good
Figure 7-4 The Hershey-Chase experiment.
If DNA is the hereditary material then:
1) Cells must be able to replicate DNA. How? Mold? Template?
2) Carries hereditary information. New processes needed to explain these.
3) Transfer information so as to control a cell's activity. How can a molecule carry info?
4) Must be able to change (mutate). Carry one information type then another? How?
What were the basic facts that were known to Watson and Crick at the time of their
discovery.
DNA has a double-helix structure
J. Watson & F. Crick (Nobel Prize in 1962; Watson, Crick, & Wilkins) 1953 Cambridge
Univ.
No experiments - work was theoretical
- few facts -> synthesized the correct answer
Few facts where: Biochemists -> DNA composed of 4 bases + sugar
deoxyribose (form nucleotides) + phosphoric acid
X-ray diffraction data (from Rosalind Franklin and
Maurice Wilkins) suggested DNA was:
- 2 long strands twisted around one another
- thickness of ~ 20 Ångstrom
Chargaff (1950) ->
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Lecture 1; 2007
Section B2; Good
- amounts of A = T , G=C
- T and C small molecules
- A and G are larger molecules
Problem solved
Put it all together ->
Double Helix Model Fig 7-9, -10 –11 (8th)
- 2 chains of nucleotides held together by phosphodiester bonds (phosphate)
- 2 chains held together by hydrogen bonds in an antiparallel manner
Accounts for:
1) X-ray diffraction data
2) Chargaff"s ratios A+G = (C+T) A=T G=C
Implications of the structure
Obvious explanation of 3 of 4 previous requirements of DNA being the genetic material
are inherent in the structrure:
1) Replicate DNA with A:T G:C pairs - complementary pair -> exact duplication
2) Information carried as the sequence of bases How?-> details determined later
3) If sequence is the information then changes in sequence
-> Changes in information -> mutations
I expect you to know that A,G = purines and CT = pyrimidines, why they pair well (size,
electrostatic interactions) and that G:C pair more tightly that A:T.
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