Download HGP - eduBuzz.org

Applications of DNA
technology
Human Genome Project
Human Therapeutics
Forensic Uses
Agriculture
History



Late 1980’s idea was proposed
Predicted it would take 15 years
Cost about $200 million per year





$1 per base pair
Officially began in 1990
26 June 2000 joint announcement from Blair and
Clinton ‘the draft complete’
Joint publication in Nature and Science 12 Feb 2001
14 Apr 2003 – The finished human genome
Why?

‘If I were to study the carburettor of my car
engine, even in exquisite detail, I would still have
no idea about the overall function of the engine,
much less the entire car. To understand what an
engine is for, and how it works, I’d need to study
the whole thing – I’d need to place the
carburettor in context, as one function part
among many. The same is true of genes…..’
James Watson
Method

Genetic Mapping
Identifies relative positions of genes
 E.g. Gene 2 lies between genes 1 and 3


Physical Mapping
Absolute positions of genes on chromosomes
 E.g. Gene 2 is 1 million bp from gene 1


DNA sequencing

Actual ATCG combinations
Polymerase Chain Reaction


Critical to the HGP was the ability to create large
amounts of DNA for sequencing.
PCR is a process to amplify DNA




DNA is heated to 95oC
The DNA is denatured causing the two strands to separate
A primer (short length of DNA) binds (or anneals) to the
template strands [after the solution is cooled]
Complementary DNA strands form [through the action of
DNA polymerase]
Genetic Mapping


Genetic mapping utilises cross over frequencies
between known genetic markers.
A genetic marker is any sequence of genome
that shows difference between individuals. It
could be
A gene
 A Microsatellite or Short Tandem Repeat

Microsatellites (STRs)



Repeating sequences 2 – 4 nucleotide bases found in
mostly in introns (‘junk’ DNA)
The number of repeats varies from person to person,
but follow patterns of Mendalian inheritance
These sequences can be identified using probes. A
probe is



A short, single stranded sequence of DNA
Complementary to DNA base sequence
‘Tagged’ – radioactively or using fluorescent dye
Recombinance / Cross Over
Frequency


What do you remember about linked genes from
Higher.
What information do recombinant individuals
give us about the relative position of genes?
Physical Mapping

Use of restriction enzymes
Restriction enzymes belong to the group
‘endonucleases’ or nucleases.
 They cut DNA at specific sequences.
 Chromosomes can therefore be cut into sequences
of different lengths.
 By using combinations of restriction enzymes and
working out the size of the fragments, a pattern of
recognition sites in the DNA can be pieced together.

Gel electrophoresis




Uses physical properties of DNA (size and charge) to
separate molecules
Gel electrophoresis involves running an electric current
through an agarose gel. DNA is loaded into wells at
the negative end.
DNA is repelled, and moves through the agarose gel at
different speeds depending on the size of a fragment.
A Molecular weight marker (of known fragment size) is
also used. Samples can be compared to this marker to
work out their size.
Physical Mapping cont…
M = molecular weight
marker
1 = undigested sample
2 = digested with NotI
3 = digested with BamHI
4 = digested with NotI and
BamHI
5 = something totally
different
Physical Mapping cont…
Physical Mapping cont….
DNA Sequencing

Sanger method (aka dideoxy chain-termination
method)

An unknown DNA template strand is replicated using a
primer (to initiate replication), DNA polymerase,
nucleotide bases and one of 4 dideoxy nucleotide bases
(ddA, ddG, ddC and ddT)
DNA sequencing cont…

If a ddNTP is inserted instead of a normal
nucleotide DNA replication stops at that point
(chain termination)

When this experiment is repeated with other dNTPs
DNA molecules differing in length by 1 bp are
created.
DNA sequencing cont..

These are then run on an electrophoresis gel
Note: The smallest pieces at the start of the sequence are at the
bottom. Therefore the sequence is read bottom to top
DNA sequencing cont…
Comparing Genomes




Other genomes have been sequenced in
conjunction with the human genome, and have
continued.
There are a large number of similarities between
genomes.
Genes found in a wide range of organisms are
known as homologous genes.
These are used to study function of genes.
Learning Activities






Look at the arrangements document to clarify
what information is required.
Read DART pg 73 – 81.
Read the Monograph pg 67 – 79
Scholar – 8
Internet research
Worksheets