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(1) DNA Profiling
DNA profiling is being used as part of an international research project to identify twenty
thousand marine fish species by the year 2010. Scientists will identify the sequence of bases
in a specific region of mitochondrial DNA which does not show much variation within species,
but between species is very different. The DNA is sourced from muscle tissue using standard
DNA extraction techniques.
(a)
Describe what DNA profiling is.
(b) PCR and gel electrophoresis are two of the techniques used in DNA profiling. For each
of these two techniques discuss BOTH, how the technique is carried out and why the
technique is used.
(c)
Discuss the human need or demand for DNA profiling technology in relation to
identifying marine fish species.
(2) Genome Analysis
“Sheep genomics” is a trans-Tasman partnership involving Australian and NZ research
organisations. The project will ultimately lead to sequencing of the sheep genome. This will
allow a virtual sheep genome map to be created, according to Australian researcher Dr Brian
Dalrymple. Around the world genome analysis is currently being carried out on several species.
(a)
Describe what genome analysis is.
(b) Two techniques used in genome analysis are the development of DNA sequencing and
DNA chips. Discuss how and why BOTH DNA sequencing and DNA chips are used in
genome analysis.
(c)
Use the above examples of genome analysis to discuss why people are spending huge
amounts of time and money to carry out this application in sheep.
(3) Transgenesis
Chymosin is an enzyme that digests milk proteins and can be used to clot milk into curds.
Traditionally Chymosin was obtained from the stomach secretions of suckling calves less than
10 days old but is now produced in an application of Transgenesis by using ‘transgenic bacteria’ –
these are bacteria which have had the chymosin gene added to their DNA.
(a)
Describe what Transgenesis is.
(b) Discuss how restriction enzymes and DNA ligase are used produce transgenic bacteria,
such as the chymosin producing bacteria.
(c)
Discuss why producing chymosin has changed from extraction from stomach secretions
of suckling claves less than 10 days old to the use of transgenic bacteria.
(4) Gene Cloning
Since Banting and Best discovered the hormone, insulin in 1921, diabetic patients, whose
elevated sugar levels are due to impaired insulin production, have been treated with insulin
derived from the pancreas glands of abattoir animals. The hormone, produced and secreted by
the cells of the pancreas regulates the use and storage of food, particularly carbohydrates.
Although bovine and porcine insulin are similar to human insulin, their composition is slightly
different. Consequently, a number of patients' immune systems produce antibodies against it,
neutralising its actions and resulting in inflammatory responses at injection sites. Added to
these adverse effects of bovine and porcine insulin, were fears of long term complications
ensuing from the regular injection of a foreign substance, as well as a projected decline in the
production of animal derived insulin. These factors led researchers to consider synthesising
Humulin by inserting the insulin gene into a suitable vector, the E. coli bacterial cell, to produce
an insulin that is chemically identical to its naturally produced counterpart through Gene
Cloning.
(a)
Describe what Gene Cloning is in relation to the example given.
(b) Restriction enzymes and ligation are two key techniques used in Gene Cloning. Discuss
how each technique is carried out and include an explanation as to why each technique is
used to clone the lysozyme gene using a plasmid vector.
(c)
Discuss the human needs or demands for this process and its particular usefulness. In
your discussion include potential advantages and disadvantages of this form of the
production of the enzyme.
(1) DNA Profiling - Answers
(a)
Achieved: DNA profiling is a method that uses microsatellities or STRs in DNA to identify
different species or individuals OR create patterns (ladders) from which the DNA can be
compared to ascertain relatedness of two or more samples or to identify specific features
when compared to a known sample.
(b)
Achieved: describes how OR why the TWO techniques are used
e.g. PCR is used to increase the amount of DNA you have. PCR can change a very small sample
of DNA into a large sample very quickly.
e.g. Gel electrophoresis separates DNA fragments by their size.
Merit: explains how or why ONE of the two techniques are used
e.g. WHY - PCR is used to amplify the DNA sample so that many tests can be taken from the
original sample of tissue taken from the victim.
HOW – Raising the temperature breaks the hydrogen bonds between the two strands
because they are relatively weak bonds. Lowering the temperature anneals the primers to
the DNA strand, raising the temperature again allows the DNA Polymerase to add
nucleotides to the primers.
e.g. WHY - Gel Electrophoresis is used because the technique separates the DNA fragments
by their size making it possible to analyse the pattern produced by fish species. Thereby
identifying differences between species and recognising the pattern produced by each
species.
HOW – Electric field through agar gel separates fragments according to size. DNA is
negatively charged so moves towards the positive electrode. The smaller fragments are able
to move faster because of their size so in a set time they move further from the starting
point. The spread out pattern produced by each species is different because the length of
the microsatellite is different so this gives a way of recognising each of the different fish
species.
Excellence: discusses how AND why ONE of the two techniques is used.
(c)
Achieved: describes human need or demand.
e.g. The use of DNA profiling enables fish species to be accurately identified OR this allows
biodiversity / conservation issues to be addressed OR evolutionary links identified OR data
on fish species will also enable fish quotas / stocks to be ascertained.
Merit: explains human need or demand.
e.g. DNA profiling is faster and more accurate than any other method of identification so
that data on fish species can enable fish quotas / stocks to be ascertained to ensure they
are sustainable.
Excellence: discusses human need or demand
e.g. This use of DNA profiling is justified as it enables fish species to be identified faster
and more accurately than any other method. This allows biodiversity and conservation issues
to be addressed more accurately, as data on each fish species is more accurate. Without the
use of DNA profiling different species of fish that look similar may have been misidentified
and this would have made it appear that there were more of a species present than was
actually the case. When the stocks of a fish species are able to be quickly and accurately
determined it helps to ensure that appropriate management strategies are implemented to
maintain a sustainable catch.
(2) Genome Analysis - Answers
(a)
Achieved: Genome analysis is about finding out the order of bases in the DNA as well as
finding the location of genes and the parts of the DNA that controls the activity of the
genes.
(b)
Achieved: describes how OR why the TWO techniques are used
e.g. DNA sequencing determines the order of bases of the genome
e.g. DNA chips are used as a tool to analyse the presence or absence of a gene/sequence
of bases in the genome.
Merit: explains how or why ONE of the two techniques are used
e.g. WHY – DNA sequencing – by determining the exact sequence of the bases, scientists
can identify the location of the genes and their known base sequence and use this
information to screen for mutations and advantageous genes.
HOW – DNA sequencing – mixture of target DNA, excess nucleotides, trace labelled
nucleotide, DNA polymerase, fragments of assorted length synthesised based on
incorporation of nucleotide analogues, fragments separated by electrophoresis, terminal
nucleotide established through fluorescence
e.g. WHY – DNA chips can be used to determine if a gene of interest is present or absent
very quickly, by having the complementary sequence on the DNA chip and adding an
individuals DNA (single-stranded). The fluorescent tag indicates to the scientists if the
gene of interest is present or not.
HOW – DNA chips – a single stranded DNA or mRNA is attached onto a grid that is
divided up. The target of single-stranded DNA or mRNA, is then introduced to the grip,
and once it locates a complementary sequence, base pairing occurs. A fluorescent tag
lights-up/indicates when base-pairing has occurred.
Excellence: discusses how AND why ONE of the two techniques is used.
(c)
Achieved: describes human need or demand.
e.g. Sheep genome analysis enables scientists to identify sheep with the genes for
greater productivity.
Merit: explains human need or demand.
e.g. Sheep genome analysis enables scientists to identify sheep with the genes for
greater productivity. This information can then be used in breeding programmes to
produce whole flocks that are carrying the genes for greater productivity.
Excellence: discusses human need or demand
e.g. Sheep genome analysis enables scientists to identify sheep with the genes for
greater productivity. This information can then be used in breeding programmes to
produce whole flocks that are carrying the genes for greater productivity.
The identification of specific genes allows the improved features to be established much
more quickly in a breeding programme than would be possible without the genome analysis
information.
(3) Transgenesis - Answers
(a)
Achieved: Transgenesis is when genetic material (DNA) is transferred from one
organisms to another of a different species. It allows for direct modification of an
organisms genome.
(b)
Achieved: describes how OR why the TWO techniques are used
e.g. Restriction enzymes are used to cut the plasmid vector and to the cut the Chymosin
gene.
e.g. Ligase is used to join wanted chymosin gene to the plasmid vector.
Merit: explains how or why ONE of the two techniques are used
e.g. WHY – Restriction enzymes are used to cut the chymosin gene and the plasmid. The
same restriction enzyme must be used to cut both the gene and the plasmid as this will
produce the same sticky ends that can then be joined together by DNA ligase..
HOW – Use of specific restriction enzymes has enabled the cutting of DNA at a
recognition site specific to that enzyme. For example the restriction enzyme EcoR1 from
E coli has the recognition site GAATTC. It will produce a DNA fragment with the sticky
end AATT at one end and TTAA at the other. During ligation of the DNA into the
plasmid the AATT end will be joined to the TTAA end of the open plasmid. When both
ends are joined in again a molecule of recombinant DNA is produced.
e.g. WHY – DNA ligase joins together sticky ends that have been created through the
use of restriction enzymes.
HOW – Weak bonds between the wanted chymosin gene and the plasmid vector form
when their sticky ends connect. DNA ligase moves along the two to make more
permanent, stronger hydrogen bonds between the bases.
Excellence: discusses how AND why ONE of the two techniques is used.
(c)
Achieved: describes human need or demand.
e.g. Chymosin can be produced on demand in the quantities required.
Merit: explains human need or demand.
e.g. Chymosin can be produced on demand in the quantities required at any time of the
year instead of having to wait until you have calves that are less than 10 days old. This
means that it is more readily available throughout the year.
Excellence: discusses human need or demand
e.g. Chymosin can be produced on demand in the quantities required at any time of the
year instead of having to wait until you have calves that are less than 10 days old. When
compared to calves transgenic bacteria are easily housed and the culture easily
maintained in vats that have the right conditions for bacterial growth and reproduction.
All the materials the bacteria need to produce chymosin are supplied and the
temperature can be held at the optimum level so that chymosin production is very
efficient.
(4) Gene Cloning - Answers
(a)
Achieved: Gene cloning is the process that allows us to obtain a gene, insert it into a
suitable vector and then make multiple copies of the gene.
(b)
Achieved: describes how OR why the TWO techniques are used
e.g. Restriction enzymes are used to cut the plasmid vector and to the cut the Insulin
gene.
e.g. Ligase is used to join wanted Insulin gene to the plasmid vector.
Merit: explains how or why ONE of the two techniques are used
e.g. WHY – Restriction enzymes are used to cut the insulin gene and the plasmid. The
same restriction enzyme must be used to cut both the gene and the plasmid as this will
produce the same sticky ends that can then be joined together by DNA ligase..
HOW – Use of specific restriction enzymes has enabled the cutting of DNA at a
recognition site specific to that enzyme. For example the restriction enzyme EcoR1 from
E coli has the recognition site GAATTC. It will produce a DNA fragment with the sticky
end AATT at one end and TTAA at the other. During ligation of the DNA into the
plasmid the AATT end will be joined to the TTAA end of the open plasmid. When both
ends are joined in again a molecule of recombinant DNA is produced.
e.g. WHY – DNA ligase joins together sticky ends that have been created through the
use of restriction enzymes.
HOW – Weak bonds between the wanted chymosin gene and the plasmid vector form
when their sticky ends connect. DNA ligase moves along the two to make more
permanent, stronger hydrogen bonds between the bases.
Excellence: discusses how AND why ONE of the two techniques is used.
(c)
Achieved: describes human need or demand.
e.g. Insulin can be produced on demand in the quantities and chemical structure required.
Merit: explains human need or demand.
e.g. Insulin can be produced in a way that has a chemical composition identical to human
insulin and can also be produced on demand in the quantities required.
Excellence: discusses human need or demand
e.g. Insulin can be produced in a way that has a chemical composition identical to human
insulin and can also be produced on demand in the quantities required. In comparison to
insulin that used to be derived from bovine and porcine, the insulin produced through
gene cloning will trigger the production of antibodies as it’s chemical composition will not
be identified as foreign by the human immune system.