NATIONAL QUALIFICATIONS CURRICULUM SUPPORT
Biology
The Structure and Replication
of DNA
Support Materials
HIGHER
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Acknowledgement
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© Learning and Teaching Scotland 2011
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Contents
1(a)
DNA
4
1(a)(i)
The structure of DNA
6
1(a)(ii)
Organisation of DNA in prokaryotes and eukaryotes
19
1(a)(iii)
The polymerase chain reaction (PCR)
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These notes have been designed to support t eachers in delivering the content
of the new Higher Biology syllabus (2011). Possible student activities are
identified throughout. Background information has been written to provide
the necessary scientific context to the topic and often goes beyond the detail
required of the student. Key concepts have been identified throughout at the
level of detail it is felt the student will need for the new Higher Biology
syllabus.
1(a) DNA
Curriculum content notes
DNA encodes hereditary information in a chemical language. All cells store
their genetic information in the base sequence of DNA. The genotype is
determined by the sequence of bases.
Key concepts




DNA is inherited.
DNA is the genetic material of living things.
DNA is located within the nucleus of all cells apart from red blood cells.
DNA is a long chemical sequence and this sequence contains the
information needed for that living thing to develop, survive and pass its
genetic information on to the next generation .
 The DNA chemical sequence differs between individuals. The pattern of
this sequence is called the genotype.
Background information
Students should have met the concepts of genetic information and genotypes
previously in the topic of inheritance in Standard Grade or to some degree in
Intermediate studies. The rest of this unit will move the student on to a
biochemical understanding of the nature of inheritance through the molecule
DNA.
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Student activity 1: DNA is the instructions for all living things
To begin with, a recap on the basics may be useful: DNA/genes hold the
instructions for all living things, it is found in cells and it is the hereditary
material. A fun way to do this might be to show a simple animation that sums
up this information, for example:
http://www.genejury.biology.ed.ac.uk/genesanimation.html
http://www.genesareus.org/filmlibrary/whatgenesmeans
Student activity 2: DNA extraction
If students have not extracted DNA before then carrying out an extraction
practical would be useful at this point before their knowledge is extended to
an understanding of the molecular structure of DNA in 1(a)(ii). Carrying out
DNA extraction helps the student see that DNA exists in all living things and
that it is held within cells.
There are many extraction protocols to be found on the internet. Here are
some of the best:
1.
The Genes are Us website has lots of videos and information about
genetic disorders and gives an extraction protocol called ‘DNA
cocktail’ with teacher’s notes and student sheet. Find it by following
this link and looking in resources for 14–15-year-olds:
http://www.genesareus.org/keystage4
2.
From The University of Utah website:
http://learn.genetics.utah.edu/content/labs/extraction/howto/
3.
From The Naked Scientists website (Cambridge University):
http://www.thenakedscientists.com/HTML/content/kitchenscience/exp/h
ow-to-extract-dna-from-a-kiwi-fruit/
Student activity 3: DNA – true or false
This is a PowerPoint presentation showing true or false statements. It can be
used to go through the basics with the students and to ascertain their
understanding and gaps in knowledge. Students could use ‘show me’ boards
as a whole class or carry out a pair exercise before feeding back answers to
the class.
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1(a)(i) The structure of DNA
Curriculum content notes
The structure of a DNA nucleotide (deoxyribose sugar, phosphate and base).
Nucleotides bond to form a sugar–phosphate backbone. Base pairs (adenine,
thymine, guanine and cytosine) held by hydrogen bonds , forming a double
helix. Double-stranded anti-parallel structure with deoxyribose and phosphate
at 3′ and 5′ end of each strand.
Key concepts




DNA is composed of two polynucleotide chains.
Nucleotides consist of a sugar, phosphate and base.
Nucleotides bond to form a sugar–phosphate backbone.
The two polynucleotide chains run antiparallel , with a deoxyribose sugar
at the 3′ end and a phosphate group at the 5′ end.
 The nucleic acid bases are paired by hydrogen bonding in the centre to
form a double helix.
 Base pairing is specific, with adenine pairing with thymine and cytosine
pairing with guanine.
Prerequisite knowledge
 DNA is the genetic material.
Background information
Knowledge of the molecular structure of DNA was part of the content of the
old Higher Arrangements, and so plenty of material showing its structure
already exists. The level of detail in the new curriculum is to be the same
apart from naming the bond type (hydrogen) between bases and the term
‘anti-parallel structure’, although the nature of this structure is inherent in the
double helix, which was on the syllabus previously. Here are some good
websites for information and resources:
1.
The University of Utah has two excellent sites with high-quality
resources for teaching and learning genetics.
http://learn.genetics.utah.edu/
http://teach.genetics.utah.edu/
2.
To order a model of DNA visit this site:
http://www.molymod.com/
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3.
Nature Publishing Group
http://www.nature.com/scitable
4.
The Wellcome Trust produces high-quality material with an emphasis
on human health. In particular, they produce a fun and scientifically
rigorous booklet called The Big Picture and their January 2010 copy
focuses on genes, genomes and health. Print copies can be ordered and
a pdf is available. Very accessible and interesting for young people.
http://www.wellcome.ac.uk/Education-resources/Teaching-andeducation/index.htm
5.
Nucleic acid problem set:
http://www.biology.arizona.edu/molecular_bio/problem_sets/nucleic_ac
ids/nucleic_acids_1.html
6.
A-level biology revision site:
http://www.s-cool.co.uk/
Student activity 4
Create your own DNA using PowerPoint or cutting and pasting
Two PowerPoints are provided that can be used by students to construct the
molecular structure of DNA.
In the first PowerPoint the student drags and drops the pieces to create a
double-stranded molecule of DNA that is five nucleotides long. This can be
done by moving pieces to the coloured space surrounding the white
PowerPoint slide, copying and pasting the shapes to create enough for the
model and then dragging and dropping the shapes back on to the slide to
represent the molecular structure.
The second PowerPoint can be printed on A4 or A3 paper so that students can
cut out the pieces and use them on a table top. These could be stuck into their
books or alternatively kept in small packs (mayb e laminated) to be used
again. If they are to be used again the students could draw the model and
label it to help reinforce their learning.
Student activity 5: Make your own edible DNA double helix
Go to the University of Utah’s excellent Teach Genetics website:
http://teach.genetics.utah.edu/
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Follow the Print-And-Go™ Lesson Plan Index and you will find a wealth of
activities, including instructions on how to make a double helix out of sweets.
Student activity 6: How the structure of DNA was discovered
This activity will help students appreciate the process of scientific discovery,
develop a deep understanding of the structure and function of DNA, develop
research skills and be able to present work in the style of a scientific poster.
This activity could be used after the class have looked at the structure of
DNA in detail and is one of the suggested learning activities in the new
Higher Biology syllabus.
Learning outcomes for activity 3
1.
2.
3.
4.
5.
6.
7.
Develop an understanding of the means by which living things can pass
on information to the next generation via DNA.
Develop an appreciation that scientific discovery is in the context of a
community of scientists, where the results of one team lay the
foundations for the work of others.
The elucidation of the genetic code by Watson and Crick as an example
of drawing conclusions from a body of scientific evidence (evidence based conclusion).
Identification of the different forms by which scientific work is
presented and the advantages and disadvantages of these different
formats, including journal papers, posters and conference presentations.
Develop an understanding of the structure of DNA (level dependent on
whether this becomes before or after Student activity 2 and whether this
is emphasised during the presentation of the posters).
Development of skills in using the internet for research: selecting
appropriate information, summarising material, identifying reliable
sources.
Development of team-working skills: assigning roles, planning work.
Introduction
The elucidation of the genetic code by Watson and Crick in 1953 was the
culmination of years of scientific research by a range of scientists. The
research and events surrounding the discovery have been the topic of many
books, media stories and even a film. The story is not only one of scientific
discovery but also of personalities and intrigues. It lends itself well to
independent research and to developing a conceptual unde rstanding of the
significance, structure and function of DNA, laying down the foundations for
Unit 1 of Higher Biology. An appreciation of how short a time there has been
since the discovery of DNA and the huge amount of research that has
followed in the field never fails to amaze and is also important when
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considering the moral, ethical and social implications of this area of
biological science.
This activity will see students examine and present the results of six
individuals or groups of scientists that were all involved in the work that led
to the discovery of the structure of DNA:






Griffiths
Avery et al.
Hershey and Chase
Chargaff
Wilkins and Franklin
Watson and Crick.
Background information for teachers
This section provides the necessary background surrounding each of the
scientists’ work. This text is not intended for students and is pitched above
the content needed for Higher.
At the turn of the 20th century, scientists knew that chromosomes contained
the genetic material that enabled organisms to pass on hereditary material or
‘genes’ to the next generation. Over the next 30 or so years it became evident
that chromosomes were composed of protein, DNA and RNA , with protein
identified as the most likely candidate for the genetic material. Given
protein’s 20 amino acids, it was felt to be sufficiently complex for the huge
amount of information that would need to be carried, in contrast to the
perceived simplicity of the structure of DNA.
Over the next 50 years the work of a number of scientists would reveal that it
was in fact DNA that was the genetic material, that the nucleic acid bases of
DNA were found in set ratios, that DNA was a helical structure and finally
that it was a double helix, composed of paired bases along two sugar–
phosphate backbones, running parallel but in opposite direct ions. Let us now
look at the work of the six scientists highlighted and the contribution they
made to the elucidation of the structure of DNA.
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Fredrick Griffith – The ‘transforming principle’
A British medical officer, Fredrick Griffith, worked on
the pneumonia-causing bacteria Streptococcus
pneumonia. In 1928 he conducted a series of
experiments revealing the ability of one strain type to
‘transform’ into another strain type through a process
involving an agent that he coined the ‘transforming
principle’.
In brief, there were known to be two strains of the
bacterium: the smooth (S) and rough (R) strains, with
the S strain being highly virulent and the R strain not. For each strain there
were two types (type II and type III), each capable of mutating from one
strain to the other. For example, type IIR is capable of transforming into IIS
and vice versa. However, this change is type -specific, so type II cannot
mutate into type III and vice versa. In Griffith’s final experiment shown in
the last column in Figure 1, he infected mice with living IIR bacteria along
with heat-killed IIS cells. The mice died, showing that the living IIR bacteria
had somehow ‘transformed’ into the virulent IIIS type by interaction with the
dead IIS cells. Griffiths concluded that some agent was involved, which he
believed to be a protein, and he called this agent the ‘transforming principle’.
Figure 1: Griffith’s experiment, showing the fate of mice when infected with
different strains or strain combinations of the Streptococcus pneumonia virus. The
experiment in the final column revealed to Griffith that there is a ‘transforming
principle’, which he assumed to be protein. Reproduced from Wikimedia Common.
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Oswald Avery – DNA is the genetic material
Along with his colleagues Colin MacLeod and Maclyn
McCarty, Avery conducted a series of experiments in the
1930s and 1940s to find out what the ‘transforming
principle’ was. He found evidence suggesting that it was
DNA.
Avery and his colleagues recreated Griffith’s experiments
in a test-tube by incubating the cell extract from a lysed
IIIS cell (and thus dead cells) with that from IIR cells.
They then observed the resulting colony types that grew on
medium in a Petri dish.
Amongst the resulting colonies were those of IIIS type , showing that they had
recreated Griffith’s transformation. They therefore knew that somewhere
amongst the cell extract was the ‘transformation principle’. The key challenge
for Avery and his team then was to deduce which of the macromolecules
present was the important one: polysaccharide, protein, RNA or DNA?
Through a series of systematic experiments they used enzymes to degrade
each of the macromolecules one at a time. They discovered that it was only
when DNAse (which degrades DNA) was used that the transformation ceased
and no type IIIS colonies grew. Thus they deduced that it was in fact DNA
that was the genetic material. However, the experiments were criticised by
the scientific community, who strongly believed that it was protein and not
DNA that was the genetic material.
Alfred Hershey and Martha Chase – DNA is the genetic material
Hershey and Chase, two American
scientists, used the T2 bacteriophage, a
virus that infects bacteria, to show that
DNA was indeed the genetic material.
They knew that bacteriophages
consisted of only proteins and DNA,
and set out to explore which of these
macromolecules was the genetic
material of the virus. Bacteriophages
cannot reproduce themselves, utilising
the host bacterium’s molecular
mechanisms to produce T2 progeny. With this in mind, Hershey and Chase
developed an experiment using radioactively labelled viral protein and DNA,
to see which could be found in the host cell after infection with the T2
bacteriophage; whichever it was must be the genetic mat erial.
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1
2
Figure 2: Hershey and Chase’s experiment. Experiment 1, on the top line, shows the
results of when DNA was radioactively labelled, with radioactivity being detected
within the cell. Experiment 2 shows the results when protein was radioactively
labelled, with radioactivity being detected in the solution that contained the ‘phage
ghosts’. The results revealed that DNA was the genetic material. Reproduced from
Magnus Manske on Wikipedia commons.
Cells of the bacterium Escherichia coli were grown in media containing a
radioactive isotope, either 32 P or 35 S, and were infected with the T2 virus.
After infection and the production of progeny, Hershey and Chase were left
with two types of virus. As DNA contains phosphorus and not sulphur and
protein contains sulphur not phosphorus, one T2 type had radioactively
labelled protein, the other radioactively labelled DNA.
They infected more E.coli cells with the radioactively labelled T2
bacteriophage, one set with the DNA labelled and the other with the protein
labelled (see Figure 2). They predicted that shortly after infection the E. coli
cells would contain the genetic material of the T2 bacteriophage, before it
reproduced and developed into progeny phages. Outside the cell would be the
virus debris (called the phage ghost). For bacteria cells infected with DNAlabelled T2 virus, they detected radioactivity within the bacteria shortly after
infection. For bacteria infected with protein-labelled virus, they detected
radioactivity in the phage ghosts outside the bacteria. Thus, they concluded
that DNA was indeed the genetic material.
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Erwin Chargaff – Chargaff’s rules and the base composition of DNA
Chargaff trained as a chemist. He analysed biological cell materials at
Columbia University in New York City. During the late 1940s Chargaff
hydrolysed the DNA from a number of different organisms and found that
DNA always contained 50% purine bases and 50% pyrimidine bases.
Secondly, his analysis showed that there was always an equivalent number of
adenine and thymine bases, and of guanine and cytosine bases. Thirdly, he
found that the percentage of each base was species specific. His work was
extremely important in refuting beliefs that DNA was not complex enough to
explain biological diversity, as up to this time it was widely held that DNA
consisted of regular repeats of the four bases . If the bases were not arranged
in the same pattern across all species, there was the possibility of DNA
holding the key to the diversity of life.
Chargaff’s base composition studies were critical in Watson and Crick’s
model structure of DNA. The feelings of Chargaff about Watson and Crick’s
work makes interesting reading and would be an interesting start to a debate
about the discovery, following the completion of this task. The references at
the end will lead you to websites and books that will give insight into the
controversy.
Rosalind Franklin and Maurice Wilkins – DNA is helical and has
measured regularities in its structure
Using a technique called X-ray diffraction, Franklin and Wilkins produced
data to give them an insight into the molecular structure of DNA. The
technique involved firing beams of parallel X-rays at isolated strands of
DNA. When the X-rays collide with the atoms they diffract to an extent that
is dependent on the molecular weight and spatial arrangement of the atoms.
These diffracted X-rays are then collected on a photographic plate and the
resulting pattern analysed.
Franklin was able to deduce from these results that DNA was helical and that
there were noticeable structural regularities along its axis at 0.34 nm and 3.4
nm. Franklin’s unpublished X-ray data was shown to Watson and Crick
during their visit to Kings College, London, where both Wilkins and Franklin
worked. Two papers, one from Wilkins and one from Franklin, were
published alongside Watson and Crick’s Nature paper, describing the
experimental data they had produced.
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James Watson and Francis Crick – a model for the physical and chemical
structure of DNA
In 1953 two Cambridge scientists, James Watson and Francis Crick,
published an extremely important and now famous journal paper in Nature
entitled ‘Molecular structure of nucleic acids. A structure for deoxyribose
nucleic acid’, which described their deduction of the structure of DNA, the
genetic material of life.
Watson and Crick came to their conclusions from the experimental data of
others, particularly that of Chargaff, Wilkins and Franklin, and offered no
fresh experimental data of their own. However, the momentous breakthrough
was in piecing together all of the evidence to provide a model that fitted all
of the known data. Their work was regarded as so significant that along with
Wilkins they were awarded the Nobel Prize in Physiology or Medicine in
1962. Controversy surrounded the awards due to the omission of Chargaff
and the playing down of Franklin’s role in the discovery . However, Franklin
had unfortunately died by this point and Nobel prizes are never aw arded
posthumously.
Watson and Crick’s conclusions about the structure of DNA were as follows:
 DNA is a double helix formed from two
polynucleotide chains that are wound round in a
clockwise direction (Figure 3).
 The two polynucleotide chains are antiparallel,
meaning that they are positioned head to tail (each
chain has a 5′ end and a 3′ end and so the 5′ end of
one lies opposite the 3′ end of the other)
 The sugar–phosphate backbone, which forms from
the connection between the nucleotides, is on the
outside of the double helix. The nucleotide bases are
on the inside, perpendicular to the sugar–phosphate
backbone.
 Each base is connected to a complementary base on
the parallel chain by weak hydrogen bonds. Adenine
always pairs with thymine by two hydrogen bonds
and guanine to cytosine by three hydrogen bonds. No
other base pairings would work to give rise to this DNA structure.
 Structurally, the base pairs are 0.34 nm apart and each 360 ° degree turn of
the helix is 3.4 nm (10 base pairs). Also, as the nucleotides are unequally
spaced between the two polynucleotide strands becau se they are
antiparallel, there are unequal grooves in the helical twist.
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Further reading and other references
To find out more about each set of experiments and the story surrounding the
discovery of the structure of DNA you could refer to the following
references.
Websites
1.
Nature education site, which gives a detailed account of the story:
http://www.nature.com/scitable/topicpage/discovery -of-dna-structureand-function-watson-397
2.
Produced at Cold Spring Harbour in America , DNAI is an interactive
site for students in genetics. One of the tasks in section 1 on the menu –
‘Code’ – is about the elucidation of the structure of DNA and is written
for students to work through like a problem, giving information about
what each of the scientists in the puzzle discovered.
http://www.dnai.org/a/index.html
3.
A website dedicated to the race for DNA:
http://osulibrary.oregonstate.edu/specialcollections/coll/pauling/dna/ind
ex.html
Books
For a very readable book that describes the elucidation by Watson and Crick
from the viewpoint of Watson, and very accessible to students, the following
is highly recommended:
Watson, James (1968) The Double Helix: A Personal Account of the
Discovery of the Structure of DNA. Atheneum. ISBN 0-689-70602-2.
The discovery is also told from the viewpoint of Crick:
Crick, Francis (1988) What Mad Pursuit: A Personal View of Scientific
Discovery. Basic Books, New York. ISBN 0-465-09137-7.
and Franklin:
Maddox, Brenda (2002) Rosalind Franklin: the dark lady of DNA.
HarperCollins, New York. ISBN 0-393-32044-8.
and Wilkins:
Wilkins, Maurice (2003) The Third Man of the Double Helix: The
Autobiography. Oxford, Oxford University Press. ISBN 0-19-860665-6.
A film produced for TV’s Horizon programme in 1987, called ‘Life Story’,
chronicles the story of Watson and Crick, who race to find the structure of
DNA before Linus Pauling, Maurice Wilkins or Rosalind Franklin. The drama
was directed by Mick Jackson and has been released in various guises: Race
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for the Double Helix and The Double Helix. It can be tricky to get hold of but
your local library might have a copy or a Google search will help reveal
sources and YouTube snippets.
Possible lesson starters
1.
To encourage the class to start thinking about the structure of DNA, the
lesson could begin by examining the biological problem that scientists
were facing at the beginning of the century: what is the genetic material
of living things composed of? Scientists knew that living things
somehow passed on information but didn’t know what did this or how it
was done.
In pairs, students could consider three criteria that this ‘material’ would
have to fulfil.
Answers:
 be able to store the information to allow an organism to develop and
reproduce
 be able to replicate this information accurately
 be able to be ‘passed on’ to offspring
 be capable of change or difference to account for the variety of
living things that we see
2.
An extract of the film ‘Life Story’ (see references above).
Student activity 7
Discovering the structure of DNA
This word document lays out the task for the students.
If this is the first session where students are being asked to present findings
as a scientific poster, an introduction to the reporting of scientific results will
help students put their task into the context of real-world science. Students
could be presented with some example journals, for example Nature, Nature
Genetics or Science, to show how scientists produce papers of their work for
the scientific community. Also, highlight the use of posters and presentation s
by scientists at scientific conferences, for sharing with the community.
Students could discuss the merits of each type or presentation, including
points such as:
 peer review in journals, to ensure credibility
 speed of results to communication (speed fastest with presentation and
poster, less for journals and even slower for books)
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 option to present incomplete findings in posters and presentations and
provoke discussion
 level of detail in each format (posters are more of a summary, which
means many can be read relatively quickly, compared to journal papers,
which provide much more detail).
To enhance this session, a scientist could be asked to come into the class to
discuss how they approach these tasks and show papers/posters and
presentations they have developed as a scientist. Scientists can be contacted
through various schemes that run to assist their placement in schools, for
example STEM Ambassadors:
http://www.stemnet.org.uk/content/stem-ambassadors
and Researchers in Residence:
http://www.researchersinresidence.ac.uk
Presentation of posters
The posters could be displayed as they would be in a scientific poster session,
where the posters are displayed and people get the chance to wander round
reading each other’s work. The students could do this and jot down a question
they would like to ask about each poster. Once everyone has had time to look
at the posters each student gets 5 minutes to present their poster, followed by
a question and answer session.
Student activity 8: Timeline of events
Following the presentations the class could create a timeline with the posters
to demonstrate the order of events leading up to the eluci dation of the genetic
code by Watson and Crick. A discussion could take place about the roles and
relevance of each piece of research.
Timeline sequence:
1.
2.
3.
4.
5.
6.
Griffiths
Avery et al.
Hershey and Chase
Chargaff
Franklin and Wilkins
Watson and Crick
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Extension tasks
1.
Students could produce a short video about the scientific work they
investigated rather than a poster.
2.
Investigation of the controversy surrounding the relative weight given
to the scientists who contributed evidence towards the elucidation of the
DNA structure. Greater weight was given to certain scientists notably
Watson, Crick and Wilkins who received the Nobel prize. However, it
is argued a number of other scientists which are listed above played a
significant role. Teams could prepare a persuasive presentation as to
why the other scientists should have been given greater recognition for
their contribution to the discovery of the structure of DNA.
3.
For a creative option, students could act out the discoveries using role
play and factual information they have gathered.
Links with other areas of Higher Biology
 The experiments of Griffiths link with bacterial transformation in Unit 3.
3 (b) (ii) Recombinant DNA technology
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ORGANISATION OF DNA IN PROKARYOTES AND EUKARYOTES
1(a)(ii) Organisation of DNA in prokaryotes and eukaryotes
Curriculum content notes
DNA is a double-stranded molecule that can be circular or linear.
Circular chromosomal DNA and plasmids in prokaryotes. Circular plasmids
in yeast. Circular chromosome in mitochondria and chloroplasts of
eukaryotes.
The DNA found in linear chromosomes of the nucleus of eukaryotes is tightly
coiled and packaged with associated proteins .
Key concepts
 DNA exists in very long molecules that are packaged and organised in
cells.
 The organisation of DNA is different in prokaryotes and eukaryotes.
 Prokaryotes usually have a single circular chromosome.
 Eukaryotes usually have several linear chromosomes , which are packaged.
 Eukaryotic cells also contain mitochondrial DNA, and chloroplast DNA in
green plants.
 The DNA in chromosomes undergoes four stages of packaging to achieve
the most condensed state, seen during metaphase.
 DNA combines with proteins to achieve its packaged state.
Prerequisite knowledge
 DNA is the genetic material of living things.
 DNA structure.
 Difference between a prokaryote and eukaryote.
Background information
In the cells of both prokaryotes and eukaryotes DNA is organised into
structures called chromosomes. In eukaryotes the se are linear and numerous,
whereas in prokaryotes there is usually a single circular chromosome.
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Prokaryotic chromosomes
Prokaryotes usually have a single double-stranded and circular chromosome,
which carries the organism’s genetic material. Some prokaryotes, however,
have more than one chromosome, in either a circular or linear form, which
can be either a copy of the first or be composed of a different DNA sequence
and which can contain essential or non-essential genes. If the organism has a
second circular chromosome carrying a DNA sequence that is superfluous to
the existence of the organism then it is called a plasmid. In bacteria, the DNA
is packaged tightly, along with associated proteins, into a given area of the
cell called the nucleoid.
Supercoiling is the key way in which most prokaryotes manage to package the
huge length of DNA that makes up their genome into a small area in the cell.
If you imagine an elastic band being twisted you will at first get twists which
produce an ever-tighter wound band. If you continue, the elastic band will
begin to supercoil, producing bends of twisted elastic, ma king the elastic
band ever more compact. This is what happens when DNA supercoils and it is
a major feature in prokaryotes. Some supercoiling does exist in eukaryotes,
but only in small amounts. The chromosomes of eukaryotes use other
methods for compacting their DNA.
Further information about chromosomes in prokaryotes can be found at the
Nature Scitable site:
http://www.nature.com/scitable/topicpage/genome -packaging-in-prokaryotesthe-circular-chromosome-9113
Eukaryotic chromosomes
The most significant differences between the chromosomes of eukaryotes and
prokaryotes are that eukaryotes have several linear chromosomes contained
within a membrane-bound nucleus. The number of chromosomes is consistent
within species but can vary across species. So, for example, humans have a
haploid set of 23 chromosomes, whereas wheat has a haploid set of 7
chromosomes.
Eukaryotic cells also contain extra packages of DNA outwith the nucleus:
mitochondrial DNA (mtDNA) and chloroplast DNA (cpDNA). mtDNA is
found in both plants and animals, whereas chloroplast DNA is only found in
green plants and certain protists. Fundamentally, these genomes are not
inherited in a medallion fashion like the chromosomes contained within the
nucleus, but instead are inherited solely from the mother along with the other
organelles contained within the cytoplasm.
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MtDNA is often circular, double-stranded and lacking in the structural
proteins of the nuclear chromosomes, much like the chromosomes found in
prokaryotes. Their size varies enormously between species. For example, in
humans the mtDNA genome is 15,569 base pairs in size, in yeast it is 80,000
base pairs long and in some plants it can be as large as 2 million base pairs.
In the main, the mitochondrial genome codes for transfer RNAs (t RNAs),
ribosomal RNAs (rRNAs) and some subunits of proteins found within
mitochondria.
CpDNA is structured similarly to mtDNA: it is circular, double-stranded and
lacks structural proteins. Chloroplasts can contain many copies of cpDNA,
with the copy number variable between species. The size of the cpDNA
genome varies between species and can be anywhere between 80 and 600 kb.
Amongst the genes on the cpDNA lie those that code for the rRNAs, tRNAs
and some proteins required for translation, transcription an d photosynthesis.
The origin of mitochondria and chloroplasts has been the focus of some
debate amongst scientists. One of the theories , the endosymbiont theory,
proposes that both mitochondria and chloroplasts were originally free -living
bacteria that invaded primitive eukaryotic cells and that through the course of
evolution both the bacteria and the original host became depend ent to the
point where one could not live without the other. This seems a great theory to
share with students – to think that within each of our cells we harbour a onceliving bacterium that we now depend on for life, emphasising the
interconnectivity of living things.
The packaging of DNA in eukaryotic chromosomes
The level of organisation in the packaging of DNA is truly amazing. The
length of a DNA molecule, if held taut, end-to-end, in just one of your
chromosomes would measure 4 cm. If that wasn’t bewildering enough given
that cells are not nearly that big, our cells are capable of packaging this
amount of DNA into chromosomes 1.2–2 µm in length. This means that end to-end you could fit 10,000 chromosomes along the length of a fingernail. If
you take this figure and the fact that we have 46 chromosomes in each cell we
can calculate a total length of DNA in one human cell to be 1.84 m, or the
height of a 6-foot person. Considering the number of cells we have, we have
enough DNA that if put end to end it would reach the moon and back!
This next section will examine how the DNA in eukaryotic chromosomes is
packaged to achieve this feat of organisation. There are four levels of
packaging seen within cells, the highest of which is only seen during
metaphase (Figure 4).
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ORGANISATION OF DNA IN PROKARYOTES AND EUKARYOTES
Level 1: Nucleosomes
DNA in the form of a double helix is wound around histone
proteins, forming nucleosomes or what is commonly called
‘beads on a string’. The histones are positively charged
and so bind tightly to the negatively charged DNA. The
lengths of DNA between the nucleosomes are called linker
DNA. The length of linker DNA between nucleosomes is
constant within cells but can vary between species an d
tissues. This level of organisation is seen throughout the
cell cycle, with only transient separation during replication.
The combination of proteins and DNA is called chromatin,
so the beads on a string structure shown here is a chromatin
fibre.
Level 2: Thick chromatin fibre
The length of nucleosomes then coils to form a thicker
chromatin fibre, about 30 nm thick, due to interactions
between the nucleosomes and linker DNA. This level of
packaging can be seen during interp hase.
Level 3: Looped fibres
The thick chromatin fibre then folds along a non -histone
protein scaffold, producing fibres that are now 300 nm
thick. This level of packaging can be seen during prophase.
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Level 4: More folds to make the most compacted chromosome
The chromatin folded along
the protein scaffold then folds
further to produce the
compacted chromosomes that
are seen during metaphase.
This is DNA in its most
compacted form. Note that
this image shows a metaphase
chromosome, which consists
of two chromatids following
replication.
Figure 4: Overview of the levels of packaging seen in a metaphase chromosome .
Figure 5
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ORGANISATION OF DNA IN PROKARYOTES AND EUKARYOTES
Student activity 9: Prokaryote or eukaryote?
DNA is packaged in a different way in eukaryotes and prokaryotes.
Students cut out the statements on the sheets and place them into one
of three groups: prokaryotes, eukaryotes or both , depending on which they
are true for.
Student activity 10: Organisation of DNA in prokaryotes
Students research examples of genome organisation within disease-causing
prokaryotes and record their results in a table as shown below, or by using a
different recording method (poster, leaflet, classroom display that can be
added to by each individual or group).
Organism
Organisation
Size
Disease
eg Borrelia
burgdorferi
bacterium
A single linear chromosome plus at
least 17 small linear and circular
chromosomes
0.53 Mb
Lyme disease
Student activity 11: The packaging of DNA in eukaryotic chromosomes
The PowerPoint presentation ‘The packaging of DNA in eukaryotic
chromosomes’ shows images of each level of chromosome packaging
corresponding to the background notes above.
These stages can be talked through with the class whilst they take notes of
each stage.
Student activity 12: Beads on a string
The idea of the packaging of DNA into chromosomes can be conceptual ly
difficult for students. As a class or in groups physically model the packaging
levels using beads and string.
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You will need:
 wooden beads of the same size or plastic milk tops with holes punched
through the centre, or something similar. You will need 20 per group or 80
if working as a whole class.
 A ball of string, cut to a 4 m length if working as a class (represents 4 cm
found in a chromosome) or 1 m if working in groups (or 4 m each if you
are feeling brave!)
images of packaging levels: see Beads on a String image sheet
 scissors
 vocabulary sheet: see Beads on a String vocabulary sheet.
To represent the first level of packaging – nucleosomes – thread beads at
regular intervals of 5 cm along the string, making a knot before and after each
bead to keep it in place and to also demonstrate the reduction in size of DNA
as it wraps around the histones.
Work through the rest of the stages from memory or using the beads on a
string image sheet.
Students should work in their group and explain to one another what the
terms on the vocabulary list mean. This would also provide a chance for some
teacher assessment.
At the end, from memory alone, groups could be tested by the ca lling out a
packaging level, for example thick chromatin fibres, which they have to make
as quickly as possible.
Student activity 13: Sequencing activity – Explaining the stages of
packaging of DNA in eukaryotic chromosomes
See the Word document ‘The packaging of DNA in eukaryotic
chromosomes’.
The document shows the four images used in the background information and
the PowerPoint to show the different levels of chromosome packaging, but in
the wrong order. Students should either draw these in the correct order or cut
them out and stick them in the correct order. They can then write a short
paragraph to explain what is happening at each stage.
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ORGANISATION OF DNA IN PROKARYOTES AND EUKARYOTES
Student activity 14: Know your chromosomes from your chromatid and
your chromatin
Not surprisingly, these three terms are often confused. Students could look up
the definitions in the following glossary of genetics and share their findings
with a partner:
http://www.genome.gov/glossary
Other terms encountered when looking at the packaging of DNA in
chromosomes could also be researched.
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THE POLYMERASE CHAIN REACTION (PCR)
1(a)(iii) The polymerase chain reaction
Curriculum content notes
The polymerase chain reaction (PCR) is a technique for the amplification of
DNA in vitro.
In PCR, primers are complementary to specific target sequences at the two
ends of the region to be amplified.
DNA is heated to separate the strands. Cooling allows primers to bind to
target sequences. Heat-tolerant DNA polymerase then replicates the region of
DNA. Repeated cycles of heating and cooling amplify the region of DNA.
Key concepts





Small sections of DNA can be replicated in vitro using the PCR.
PCR manipulates the natural process of DNA replication.
PCR is now an automated technique widely used in many areas of
research and industry.
PCR requires template DNA, Taq polymerase, di-deoxynucleic acids with
each of the four DNA bases, Mg 2+ , primers and a buffer.
PCR involves continuous and repeated cycles of heating and cooling.
Prerequisite knowledge


The structure of DNA.
DNA replication.
Background information
This is a new topic for Higher Biology, which was previously only present on
the Advanced Higher syllabus. However, it is a fundamental and everyday
technique in many laboratories, whether used by academia, industry or
government.
The technique
PCR was developed by Kary Mullis in the mid 1980s, revolutionising
molecular biology. He received the Nobel Prize for chemistry for its
conception in 1993. The technique enables specific sections of DNA to be
amplified (replicated) in vitro, producing millions of copies from a DNA
template. Mullis developed the technique manually, and it can still be carried
out using water baths. However, the technique is now fully automated in
laboratories, using thermal cyclers no bigger than a bread machine.
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THE POLYMERASE CHAIN REACTION (PCR)
The technique manipulates the cell’s natural mechanism for replication by
using DNA polymerase and the following steps:
1.
2.
3.
4.
5.
Sample DNA is denatured by heating to give two polynucleotide chains.
Sequence-specific primers, which are small sequences of singlestranded DNA, typically of about 8–15 base pairs in length, anneal to
the DNA flanking the section of interest. One primer anneals to one
strand (forward primer), another to the other DNA strand (reverse
primer).
Polymerase begins to replicate the DNA section of interest using the
primers as a starter sequence. The mixture also includes :
 dideoxynucleotides of each base type to enable the formation of the
new DNA strand
 Mg 2+ , which is a polymerase cofactor
 a buffer to keep the pH stable.
The mixture now contains the original template plus the newly
amplified sections.
The cycle begins again using the original and copied DNA as templates.
As the reaction is exponential, millions of copies are produced in about
3 hours.
Figure 6 illustrates the PCR technique.
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Figure 6: The polymerase chain reaction.
Applications of PCR
PCR is used widely, for example:
1.
2.
DNA profiling/fingerprinting: PCR is used to rapidly identify
individuals. Specific regions of DNA known to vary between
individuals are amplified using fluorescently labelled primers and then
analysed using capillary gel electrophoresis. Profiling is not only used
in forensics but also in plant variety identification, paternity testing and
evolutionary biology.
Disease diagnosis: DNA sequences that are known to indicate certain
genetic disorders or diseases are amplified using PCR for the purposes
of diagnosis.
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THE POLYMERASE CHAIN REACTION (PCR)
3.
4.
5.
Archaeological analysis: Ancient DNA, degraded over the years, can
be amplified and used in archaeological, paleontologica l and
evolutionary research.
Population studies: Analysis of human or other species’ population
genetics can be rapidly performed using PCR analysis.
Sequencing: DNA sequence analysis previously took place following
lengthy cloning experiments, which have now been replaced by PCR.
For further information about PCR refer to the following website:
http://www.dnalc.org/resources/animations/pcr.html
A song about PCR can be seen here:
http://www.youtube.com/watch?v=x5yPkxCLads
And there is a PCR rap here:
http://www.youtube.com/watch?v=oCRJ4r0RDC4&feature=related
Practical work
A case study has been developed that enables students to experience PCR in
practice.
Student activity 15
See the PowerPoint presentation ‘The Polymerase Chain Reaction
This PowerPoint presentation takes students through the process of PCR
using a real-life, contemporary scientific context. Johanna Neilson is a PhD
student in evolutionary biology who works at the University of Edinburgh
and the University of Cambridge. She studies meerkats . The PowerPoint takes
you through the steps she would take to identify the parent of a meerkat baby
using real data.
The PowerPoint also presents students with questions throughout. These can
be tackled individually, in pairs or as a class. The activity could be broken up
into stages or used only in part. It includes animated slides of the PCR steps.
For more information about the meerkat project that Johanna works on see:
http://www.kalahari-meerkats.com
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