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Lecture 5 The chemical nature of the Gene

CAMPBELL BIOLOGY

Chapter 10: The Molecular Biology of the Gene

Some elegant experiments that provided insights into fundamental concepts in genetics

(1) Evidence that Genes are located on Chromosomes

1902 – McClung – a particular chromosome (X) determines sex in insects (XO = male; XX = female)

1903 – Sutton and Boveri – chromosomes behave just like the ‘unit factors’ described by Mendel

1910 – Morgan – the “white” eye color gene of Drosophila is located on the X-chromosome

- there are many other X-linked genes

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(2) But what are Genes made of?

1869 – Meischer - he separated nuclei from cells

- examined their chemical composition - extracted a phosphorus-rich polymer

called nuclein - this was actually DNA

1912 – Feulgen - developed stains for detection of proteins

- showed that the nucleus and chromosomes contained proteins as well as DNA

(3) Problem: If chromosomes contain both DNA and proteins, which one is the genetic material (genes)?

Arguments in favour of genes being made of proteins:

•  Proteins are sufficiently complex to store genetic information but DNA is not

•  Proteins - contain 20 different subunits (amino acids) and

•  There are many different types of proteins

•  DNA - contains only 4 different subunits

•  The complexity found in proteins was considered sufficient to specify genetic complexity

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Arguments in favour of genes being made of DNA

•  All cells of a given species contain a constant amount of DNA but the types and amounts of proteins differ in different cells

•  The amount of DNA doubles in every cell just before it divides and an exactly equal amount is distributed to the two daughter cells

•  Gametes contain half the number of chromosomes and half the amount of DNA

•  Therefore, although DNA is a very simple molecule, it behaves in the way genetic material would be expected to behave

During the 1940’s the widely accepted conclusion was:

  Genes are likely to be made of protein   DNA merely provides a structural framework in

chromosomes

Conclusive experimental evidence that genes are made of DNA came from studies of: (a) bacterial transformation

and (b) virus replication

Bacterial Transformation

1928  Griffith Streptococcus pneumoniae: two types

S strains - virulent:caused lethal pneumonia in mice - can be isolated from blood of dead mice

- bacterium has a polysaccharide coat - forms large, slimy colonies on agar plates

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R strains - non-virulent: doesn’t kill mice - doesn’t make a polysaccharide coat - grows as small, rough colonies on agar plates

Griffith’s experiments: inject mice with the different strain types observe effect on mice and analyse their blood for bacteria

Strain injected Effect Blood analysis Type II R none dead no bacteria present Type III S all dead Type III S present

killed Type III S none dead no bacteria present

killed Type III S + Type IIR all dead Type III S present

Pretty amazing idea to decide to test this last category!

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Explanation: Bacterial Transformation

1.  The killed Type IIIS did not come back to life!

2.  Genetic material contained in the Type IIIS cells has leaked out and entered the living Type IIR cells

3.  The genetic material has converted (or transformed) cells of the Type IIR strain into Type III S cells

4.  The genetic material from the Type III S contains the information for making the polysaccharide coat

required for virulence

5. Griffith didn’t realise this!

What is the transforming substance? Griffith’s discovery of bacterial transformation provided a way of identifying what chemical substance in the bacterium contained the genetic information i.e. the genes themselves

1944 Avery, McLeod and McCarthy Proved that the transforming substance = DNA

1.  Made a soluble cell extract from Type IIIS cell cultures 2.  Tested the cell extract for its ability to transform Type II R into Type IIIS.

Before testing for transformation 3. Treated the extract with different enzymes i.e.

(a) one that destroys proteins = protease or (b) one that destroys RNA = RNAase or (c) one that destroys DNA = DNAase

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Transformation Results

Transformation Transforming of TypeII R ? Substance

A.  S extract alone YES ?

B.  S extract + protease YES NOT PROTEIN

C.  S extract + RNAase YES NOT RNA

D.  S extract + DNAase NO must be DNA

www.visionlearning.com/library/modules/mid149/Image/VLObject-3756-080922120939.jpg

Avery, McLeod and McCarthy

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1953  A second experiment confirms that DNA = The Genetic material

The Hershey-Chase experiment:

  Used a virus that infects and reproduces inside a bacterial cell = phage

  The virus is made of 50% DNA and 50% protein

  Question: which substance contains the genetic information?

DNA contains phosphorus ( 32P) but no sulphur – can “label” the DNA with radioactive 32P

Protein contains sulphur ( 35S) but no phosphorus can “label” protein with radioactive S

Alfred Hershey and Martha Chase

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Experiment 1: The protein coat of the phage is labelled with 35S The DNA is not labelled.

The protein coat stays on the surface of the bacterium and can be knocked off. The progeny phage contain no 35S

Experiment 2: The DNA of the phage is labelled with 32P The protein is not labelled.

The phage injects its DNA into the bacterial cell. When the cells are pelleted, the 32P is in the pellet. The progeny phage contain 32P