Model Organisms & Tools of Cell Biology

Lecture 3

Autumn 2007

Learning the Unknown

You are a car mechanic Would you rather know a little bit about

the working of every car every constructed…

… or everything about a representative one from each category?

Science is no different!

What is a Model Organism? Many aspects of biology are similar in most or all

organisms It is much easier to study particular aspects in

particular organisms - for instance, genetics is easier in small organisms that breed quickly, and very difficult in humans!

The most popular model organisms have strong advantages for experimental research

They become even more useful when other scientists have already worked on them, discovering techniques, genes and other useful information.

How many are there? Many (about 80) Mouse, rat, zebra fish, viruses, chicken,

dog, hamster, slime mould, maize, tetrahymena, etc.

Many scientists have worked on all these over the years, and shared information extensively

Which are the main ones?

1) E. coli (bacterium) 2) Saccharomyces cerevisiae (yeast) 3) Arabidopsis thaliana (weed) 4) Drosophila melanogaster (fruit fly) 5) Mus musculus (mouse) 6) Homo sapiens (Man)

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Arabidopsis thaliana (mustard plant)

This is now the main model plant system for genetics.

Its small genome, and the recent application of classical genetics has put it far ahead of other models of agricultural importance (tomato, tobacco, corn etc.)

It's genome has been fully sequenced.

01_34_Drosophila.jpgDrosophila sp. ‘Fruit Fly’Usually the species Drosophila melanogaster - Easily raised in lab, rapid generations, mutations easily induced, many observable mutations.Many clues to development and genetics

01_38_C.elegans.jpg

Caenorhabditis elegansCaenorhabditis elegans, , a nematodea nematode(Usually called just (Usually called just C. elegans)C. elegans)-an excellent model for understanding the an excellent model for understanding the genetic control of development and physiology.genetic control of development and physiology.-C. elegansC. elegans was the first multicellular organism was the first multicellular organism whose genome was completely sequencedwhose genome was completely sequenced-First to show fixed cell count in bodyFirst to show fixed cell count in body-Gave important clues on programmed cell Gave important clues on programmed cell deathdeath

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Saccharomyces cerevisiae, baker's yeast or budding yeast (used in brewing and baking)

Early studies on this enabled us to get a great grasp on the cell cycle

Humans

• Regardless of how thoroughly we may understand other animal systems, sometimes there is no alternative but to study humans directly - i.e. breast cancers• The human animal is the most medically analyzed and documented of any species.• We have now completely sequenced our own genome too• Over the next decade or so we will understand more about our biology than ever before!

Where has knowledge of cells come from? Can we see cells? Yes and no, most are too small, but some we

can see easily - the egg of a chicken is a large single cell

What are ‘The tools of Cell Biology’? How big is the average egg? Lets put it in perspective…

Size perspective jpg1X 10X -tissues

100X - cells

1000X - organelles 10,000X - organelles

100,000X - proteins 1,000,000X - atoms

Microscopes - aid us in seeing

First cells seen by Robert Hook, who made the first compound microscope

He looked at cork cells The detail was lacking and limited by the

quality of the instrument Light microscopes are now much better

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are needed to see this picture.Light Microscopes Many types existMany types exist Light is either reflected from the specimen (Light is either reflected from the specimen (Oblique Oblique

illumination)illumination) or transmitted through the specimen ( or transmitted through the specimen (Bright Bright field optical microscopy)field optical microscopy)

How small can we see? How small can we see? We can see all cellsWe can see all cells.. The lens are so good now that the limiting factor is now The lens are so good now that the limiting factor is now

the the wavelength of lightwavelength of light itself. itself. The wavelength of the visible light used in optical microscopes The wavelength of the visible light used in optical microscopes

is between 400 and 700 nanometers (nm). is between 400 and 700 nanometers (nm). The resolving powers of high-quality light microscopes are The resolving powers of high-quality light microscopes are

limited by the wavelength of imaging light to about limited by the wavelength of imaging light to about 200 200 nanometersnanometers

Electron Microscopes Sample placed in vacuum - thus dead Four main types all using electron streams

instead of light waves; Transmission Electron Microscope (TEM) Scanning Electron Microscope (SEM) Reflection Electron Microscope (REM) Scanning Transmission Electron Microscope (STEM)

Resolutions as low as 70pm have been obtained - single atoms

• Read all about these on the course web site

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Centrifugation - for analysis Allows separation of

Cells Organelles DNA and other macromolecules

Uses gravitational and centrifugal forces to separate items based on density and size - one spins things at high speeds

Many types are used in cell biology…

Differential centrifugation

This is the most common method of fractionating cells Fractionation is

the separation of the different organelles within the cell

The speed determines which size and mass (density) of material is pelleted.

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Isopycnic centrifugation

Isopycnic centrifugation or equilibrium centrifugation is a process used to isolate nucleic acids such as DNA.

Much high speeds and duration

Separates DNA based on base composition

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Sucrose gradient centrifugation

Uses a decreasing concentration of sucrose in a tube

The particles travel through the gradient until they reach the point in the gradient at which their density matches that of the surrounding sucrose.

Other commonly used tools…

Mutation analysis - an important mechanism for discovery

Any organism which is different from the norm can be a candidate for biochemical analysis

Mutations are generally the cause of such differences

By analysis of these mutations one can gain insights or deduce the biochemical basis of important pathways…

01_35_Yeast mutation.jpg

Conditional Mutation Analysis - e.g. Temperature conditionalThe organism grows at one temperature (permissive) but fails to do so at a different one (restrictive).

All the yeast colonies grow at one temperature (23C), but if one places the same cells at 35C some of the colonies fail to grow - what is wrong with the ones that fail to grow?

01_36_S. pombe rescued.jpg

Gain-of-function mutations - e.g. biochemical function

01_37_amino acid sequ.jpg

Sequence analysis has taught us that there are about 200-300 genes required for the most basic cell survival.Comparisons across species shows that many genes are common from simple bacteria to complex life forms, and that life uses similar processes

Amoeba proteus290,000,000,000 (100 times the size of a human genome)

Bufo bufo (cane toad)6,900,000,000

Homo sapiens (Man)3,000,000,000 (3 billion base pairs - 24 chromosomes - [22, X, Y])

Muntiacus muntjak vaginalis (Indian deer)2,521,500,000

Boa constrictor (snake)2,100,000,000

Rhinolophus ferrumequinum (bat)1,929,400,000

Plasmodium falciparum (malaria parasite)25,000,000

Human immunodeficiency virus type 1 (HIV)19,750

Genome sizes - a genome is the total haploid amount of DNAGenome sizes - a genome is the total haploid amount of DNA

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There is no real correlation between the genome size and complexity

Among the organisms whose genomes are sequenced, genome size does not correlate with the number of genes.

Species Size of genome Number of genesHuman

3.0 billion base pairs 25,000 ?Fruit fly (Drosophila melanogaster)

120 million base pairs 13,601Baker's yeast (Saccharomyces cerevisiae)

12 million base pairs 6, 275Worm (Caenorhabditis elegans)

97 million base pairs 19,000E. coli

4.6 million base pairs 4,403Arabidopsis (Arabidopsis thaliana)

125 million base pairs 25,000

Genome size and number of genes

Study…

Please read chapter 1 entirely, if you have not done so already

Sample questions for the exams will start appearing on the supporting web site this week!

Bye!