Unit 5 Dna

Front 1

DNA Structure:

Back 1

1) DNA is a polynucleotide.
2) Each nucleotide is made from a pentose sugar, a phosphate group and a nitrogenous base.
3) The sugar in DNA nucleotides is deoxyribose.
4) Each nucleotide has the same sugar and phosphate. The bases can vary though.

Front 2

DNA Double-Helix:

Back 2

1) DNA nucleotides join together to form polynucleotide strands.
2) The nucleotides join up between the phosphate and the sugar, creating a sugar-phosphate backbone.
3) Two DNA strands join together by hydrogen bonding between the bases.
4) There is specific base pairing (A-T, G-C).
5) The two strands wind up to form the DNA double-helix.

Front 3

Genes:

Back 3

1) Genes are sections of DNA found on chromosomes.
2) Genes code for proteins (polypeptides).
3) Proteins are made from amino acids, and the order of nucleotide bases determines the order of amino acids in a particular protein.
4) A DNA triplet is called a base triplet or codon.
5) Different sequences of bases code for different amino acids, so the base sequence is a template used to make a protein in protein synthesis.

Front 4

Protein Synthesis:

Back 4

1) DNA molecules are found in the nucleus of the cell, but the ribosomes are found in the cytoplasm.
2) DNA is too large to move out the nucleus, so a section is copied into RNA - transcription.
3) The RNA leaves the nucleus and joins with a ribosome in the cytoplasm, where it can be used to synthesise a protein - translation.

Front 5

RNA:

Back 5

1) RNA is made of nucleotides that contain one of four different bases. (A,U,C,G)
2) The sugar in RNA is a ribose sugar.
3) The nucleotides form a single polynucleotide stand (not a double strand).

Front 6

Messenger RNA (mRNA):

Back 6

1) mRNA is a single polynucleotide strand. In mRNA, groups of three adjacent bases are called codons.
2) mRNA is made in the nucleus during transcription.
3) It carries the code from the DNA in the nucleus to the cytoplasm, where it's used to make a protein during translation.

Front 7

Transfer RNA (tRNA):

Back 7

1) tRNA is a single polynucleotide strand that's folded into a clover shape.
2) Hydrogen bonds between specific base pairs hold the molecule in this shape.
3) Every tRNA molecule has a specific sequence of three bases at one end called an anticodon. They also have an amino acid binding site at the other end.
4) tRNA is found in the cytoplasm where it's involved in translation. It carries amino acids that are used to make proteins to the ribosomes.

Front 8

Transcription - Stage 1:

Back 8

1) Transcription starts when RNA polymerase attaches to the DNA double-helix at the beginning of a gene.
2) The hydrogen bonds between the two DNA strands in a gene break, separating the strands, and the DNA uncoils at this point.
3) One of the strands is then used as a template to make an mRNA copy.

Front 9

Transcription - Stage 2:

Back 9

1) The RNA polymerase lines up free RNA nucleotides alongside the template strand. Specific base pairing means that the mRNA strand ends up being a complementary copy of the DNA template strand.
2) Once the RNA nucleotides have paired up with their specific bases on the DNA strand they're joined together, forming an mRNA molecule.

Front 10

Transcription - Stage 3:

Back 10

1) The RNA polymerase moves along the DNA, separating the strands and assembling the mRNA strand.
2) The hydrogen bonds between the uncoiled strands of DNA reform once the RNA polymerase has passed by and the strands coil back into a double-helix.

Front 11

Transcription - Stage 4:

Back 11

1) When RNA polymerase reaches a particular sequence of DNA called a stop signal, it stops making mRNA and detaches from the DNA.
2) The mRNA moves out of the nucleus through a nuclear pore and attaches to a ribosome in the cytoplasm, where translation then takes place.

Front 12

Editing mRNA - Splicing:

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1) Genes in eukaryotic DNA contain sections that don't code for amino acids.
2) These sections of DNA are called introns. All the bits that do code for amino acids are called exons.
3) During transcription the introns and exons are both copied into mRNA. mRNA strands containing introns and exons are called pre-mRNA.
4) Introns are removed from pre-mRNA by splicing - introns are removed and exons joined forming mRNA strands. This takes place in the nucleus.
5) The mRNA then leaves the nucleus for the next stage of protein synthesis (translation).

Front 13

Translation - Stage 1:

Back 13

1) The mRNA attaches itself to a ribosome and tRNA molecules carry amino acids to the ribosome.
2) A tRNA molecule, with an anticodon that's complementary to the first codon on the mRNA, attaches itself to the mRNA by specific base pairing.
3) A second tRNA molecule attaches itself to the next codon on the mRNA in the same way.

Front 14

Translation - Stage 2:

Back 14

1) The two amino acids attached to the tRNA molecules are joined by a peptide bond. The first tRNA molecule moves away, leaving behind its amino acid.
2) A third tRNA molecule binds to the next codon on the mRNA. Its amino acid binds to the first two and the second tRNA molecule moves away.

Front 15

Translation - Stage 3:

Back 15

1) The process continues, producing a chain of linked amino acids (a polypeptide chain), until there's a stop signal on the mRNA molecule.
2) The polypeptide chain (protein) moves away from the ribosome and translation is complete.

Front 16

The Genetic Code:

Back 16

1) The genetic code is the sequence of codons in mRNA which code for specific amino acids.
2) The code is non-overlapping.
3) The code is degenerate - there are more possible combinations of triplets than there are amino acids. Some amino acids are coded for by more than one base triplet.
4) Some triplets are used to tell the cell when to start and stop protein production - start/stop codons.
5) The genetic code is universal - it's the same across all living things.

Front 17

Transcription Factors:

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1) These move from the cytoplasm to the nucleus.
2) In the nucleus they bind to specific DNA sites near the start of their target genes.
3) They control expression by controlling the rate of transcription.
4) Some transcriptional factors (activators) increase the rate of transcription - they help RNA polymerase to bind to the start of the target gene and activate transcription.
5) Other transcriptional factors (repressors) decrease the rate of transcription - they bind to the start of the target gene, preventing RNA polymerase from binding, stopping transcription.

Front 18

Oestrogen Affects the Transcription of Target Genes:

Back 18

1) Oestrogen can affect transcription by binding to a transcriptional factor called an oestrogen receptor.
2) The complex moves from the cytoplasm into the nucleus where it binds to specific DNA sites near the start of the target gene. It can either act as an activator, or a repressor, and this depends on the type of cell and the target gene.
3) So, the level of oestrogen in a particular cell affects the rate of transcription of target genes.

Front 19

Small Interfering RNA (siRNA) on Gene Expression:

Back 19

1) siRNA molecules are short, double stranded RNA molecules that can interfere with the expression of specific genes.
2) Their bases are complimentary to specific sections of a target gene and the mRNA that's formed from it.
3) siRNA can interfere with both the transcription and translation of genes.
4) siRNA affects translation through RNA interference.

Front 20

RNA Interference:

Back 20

1) In the cytoplasm, siRNA and associated proteins bind to the target mRNA.
2) The proteins cut up the mRNA into sections so it can no longer be translated.
3) So, the siRNA prevents the expression of the specific gene as its protein can no longer be made during translation.

Front 21

Mutations:

Back 21

1) Mutations are any changes to the base sequence of DNA.
2) They can be caused by errors during DNA replication.
3) They can also be caused by mutagenic agents.
4) The order of DNA bases in a gene determines the order of amino acids in a particular protein. If a mutation occurs in a gene, the sequence of amino acids that it codes for could be altered.

Front 22

Types of Mutation:

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1) Substitution - one base is substituted with another, e.g. ATG becomes ATT.
2) Deletion - one base is deleted, e.g. ATGCCT becomes ATCCT.

Front 23

Mutations Affecting the Order of Amino Acids:

Back 23

1) The degenerate nature of the genetic code means some amino acids are coded for by more than one DNA codon. This means that not all substitution will result in a change to the amino acid sequence.
2) However, deletions will always lead to changes in the amino acid sequence, as it will change the number of bases present, which will cause a shift in all the codons after it.

Front 24

Mutagenic Agents:

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1) Mutations occur spontaneously, but some things can cause an increase in the rate of mutations.
2) Ultraviolet radiation, ionising radiation, and some chemicals and viruses are examples of mutagenic agents.

Front 25

How Mutagenic Agents Work:

Back 25

They can increase the rate of mutations by:
1) Acting as a base - chemicals called base analogs can substitute for a base during DNA replication, changing the base sequence in the new DNA.
2) Altering bases - some chemicals can delete or alter bases.
3) Changing the structure of DNA - some types of radiation can change the structure of DNA, which causes problems during DNA replication.

Front 26

Genetic Disorders - Hereditary Mutations:

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1) Some mutations can cause genetic disorders - inherited disorders caused by abnormal genes or chromosomes, e.g. cystic fibrosis.
2) Some mutations can increase the likelihood of developing certain cancers.
3) If a gamete containing a mutation for a genetic disorder or certain cancer is fertilised, the mutation will be present in the fetus - passed on to offspring.

Front 27

Acquired Mutations:

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1) Mutations that occur in individual cells after fertilisation (e.g. in adulthood) are called acquired mutations.
2) If these mutations occur in the genes controlling the rate of cell division, it can cause a tumour. Tumours that invade and destroy surrounding tissue are called cancers.
3) There are two types of gene that control cell division - tumour suppressor genes and proto-oncogenes. Mutations in these genes can cause cancer.

Front 28

Tumour Suppressor Genes:

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1) Tumour suppressor genes can be inactivated if a mutation occurs in the DNA sequence.
2) When functioning normally, tumour suppressor genes slow cell division by producing proteins that stop cells dividing or cause them to self-destruct (apoptosis).
3) If a mutation occurs in a tumour suppressor gene, the protein isn't produced. The cells divide uncontrollably resulting in a tumour.

Front 29

Proto-Oncogenes:

Back 29

1) The effect of a proto-oncogene can be increased if a mutation occurs in the DNA sequence. A mutated proto-oncogene is called an oncogene.
2) When functioning normally, proto-oncogenes stimulate cell division by producing proteins that make cells divide.
3) If a mutation occurs in a proto-oncogene, the gene can become overactive. This stimulates the cells to divide uncontrollable resulting in a tumour.

Front 30

Knowing the Mutation:

Back 30

1) Cancer and most genetic disorders are caused by mutations.
2) Knowing whether a disorder is caused by an acquired or inherited mutation affects the prevention and diagnosis of the disorder.
3) Identifying the specific mutation that causes a disorder in an individual affects the prevention, diagnosis and treatment.

Front 31

Stem Cells:

Back 31

1) All specialised cell types originally came from stem cells. They are unspecialised cells that can divide into other types of cells, which then become specialised.
2) Stems cells are found in the embryo and in some adult tissues (e.g. in bone marrow).
3) Stem cells that can develop into any type of body cell in an organism are called totipotent cells, and these are only present in humans in the early life of an embryo. After this point they begin to lose their ability to specialise into all types of cells, narrowing the specialisation.

Front 32

Totipotent Stem Cells in Plants:

Back 32

1) Mature plants also have stem cells - they're found in areas where the plant is growing (e.g. roots/shoots).
2) All stem cells in plants are totipotent - they can mature into any cell type.
3) This means they can be used to grow plant organs (e.g. roots) or whole new plants in vitro. Growing plant tissues artificially is called tissue culture.

Front 33

Stem Cell Specialisation - Gene Expression:

Back 33

1) Totipotent stem cells all contain the same genes - but during development not all of them are expressed
2) Under the right conditions, some genes are expressed and others are switched off.
3) mRNA is only transcribed from specific genes, and then translated into proteins.
4) These proteins modify the cell, determining structure and control cell processes etc., causing them to become specialised, which is difficult to reverse, making it remain specialised.

Front 34

Tissue Culture:

Back 34

1) A single totipotent stem cell is taken from a growing area on a plant (e.g. a root or shoot).
2) The cell is placed in some growth medium that contains nutrients and growth factors. The growth medium is sterile, so microorganisms can't compete.
3) The plant stem cell will grow and divide into a mass of unspecialised cells, which will mature into specialised cells under the right conditions.
4) The cells grow and specialise to form a plant organ or an entire plant depending on the growth factors.

Front 35

Stem Cell Therapies:

Back 35

1) Stem cells can be used to replace damaged cells.
2) Some stem cell therapies already exist for some diseases affecting the blood and immune system, e.g. bone marrow transplants for healthy blood cells, which can be used to treat leukaemia and lymphoma.
3) It has also been used to treat some genetic disorders, such as sickle-cell anaemia and severe combined immunodeficiency (SCID)

Front 36

Severe Combined Immunodeficiency (SCID):

Back 36

1) This is a genetic disorder that affects the immune system. People with SCID have a poorly functioning immune system as their white blood cells are defective, meaning they can't defend against infections, so SCID sufferers are extremely susceptible
2) Treatment with a bone marrow transplant replaces the faulty bone marrow with donor bone marrow that contains stem cells without the faulty genes that cause SCID. These then differentiate to produce functional white blood cells.

Front 37

Potential of Stem Cells on Other Diseases:

Back 37

1) Spinal cord injuries - replace damaged nerve tissue
2) Heart disease - replace damaged heart tissue.
3) Bladder conditions - grow whole bladders, which can then be implanted.
4) Organ transplants - organs could be grown.

Front 38

Benefits of Using Stem Cells in Medicine:

Back 38

1) They could save many lives - e.g. many people waiting for organ transplants die before a donor becomes available. Stem cells could be used to grow organs for those people awaiting transplants.
2) They could improve people's quality of life - e.g. stem cells could be used to replace damaged cells in the eyes of people who are blind.

Front 39

Adult Stem Cells:

Back 39

1) These are obtained from the body tissues of an adult, e.g. bone marrow.
2) They can be obtained in a relatively simple operation, with very little risk, but some discomfort.
3) Adult stem cells aren't as flexible as embryonic stem cells - they can only specialise into a limited range of cells. Scientists are trying to find ways to make adult stem cells specialise into any cell type.

Front 40

Embryonic Stem Cells:

Back 40

1) These are obtained from embryos at an early stage of development.
2) Embryos are created in a laboratory using in vitro fertilisation - egg cells fertilised outside the womb.
3) Once the embryos are approximately 4-5 days old, stem cells are removed from them and the rest of the embryo is destroyed.
4) Embryonic stem cells can develop into all types of specialised cells.

Front 41

Ethics of Stem Cell Usage:

Back 41

1) Destruction of an embryo - potential of life.
2) Some think scientists should only use adult stem cells because their production doesn't destroy an embryo. However, these are less useful.

Front 42

Gene Technology:

Back 42

Gene technology is basically all the techniques that can be used to study genes and their function, e.g.:
1) The polymerase chain reaction (PCR) - produces lots of identical copies of a specific gene.
2) In vivo gene cloning - produces lots of identical copies of a specific gene.
3) DNA probes - used to identify specific genes.

Front 43

DNA Fragment Production - Reverse Transcriptase:

Back 43

1) Many cells only contain two copies of each gene, making it difficult to obtain a DNA fragment with the target gene, but mRNA is easier to obtain.
2) The mRNA molecules can be used as templates to make lots of DNA. The enzyme reverse transcriptase makes DNA from an RNA template. The DNA produced is called complementary DNA (cDNA).
3) To do this, mRNA is isolated from cells. It's then mixed with free DNA nucleotides and reverse transcriptase, and synthesises a new strand of cDNA.

Front 44

DNA Fragment Production - Restriction Endonuclease Enzymes:

Back 44

1) These are enzymes that recognise specific palindromic sequences (recognition sequences) and cut (digest) the DNA at these places.
2) Different restriction endonucleases cut at different specific recognition sequences, because the shape is complementary to an enzyme's active site.
3) If recognition sequences are present at either side of the DNA fragment you want, you can use them to separate it from the rest of the DNA.

Front 45

Restriction Endonuclease Enzymes - Sticky Ends:

Back 45

1) The DNA sample is incubated with the specific restriction endonuclease, which cuts the DNA fragment out via a hydrolysis reaction.
2) Sometimes the cut leaves sticky ends - small tails of unpaired bases at each end of the fragment. Sticky ends can be used to bind (anneal) the DNA fragments to another piece of DNA that has sticky ends with complementary sequences.

Front 46

DNA Fragment Production - Polymerase Chain Reaction (PCR) - Stage 1:

Back 46

1) A reaction mixture is set up that contains the DNA sample, free nucleotides, primers and DNA polymerase. Primers are short pieces of DNA that are complementary to the bases at the start of the fragment you want. DNA polymerase is an enzyme that creates new DNA strands.
2) The DNA mixture is heated to 95'C to break the hydrogen bonds between the two strands of DNA.
3) The mixture is then cooled to between 50 and 65'C so that the primers can bind (anneal) to the strands.

Front 47

DNA Fragment Production - Polymerase Chain Reaction (PCR) - Stage 2:

Back 47

1) The reaction mixture is heated to 72'C so DNA polymerase can work.
2) The DNA polymerase lines up free DNA nucleotides alongside each template strand. Specific base pairing means new complementary strands are formed.

Front 48

DNA Fragment Production - Polymerase Chain Reaction (PCR) - Stage 3:

Back 48

1) Two new copies of the fragment of DNA are formed and one cycle of PCR is complete.
2) The cycle starts again, with the mixture being heated to 95'C and this time all four strands (two original and two new) are used as templates.
3) Each PCR cycle doubles the amount of DNA, e.g. 1st cycle = 4 DNA fragments, 2nd cycle = 8 DNA fragments, and so on.

Front 49

Gene Cloning:

Back 49

Gene cloning is can be done using two different techniques:
1) In vitro cloning - where the gene copies are made outside of a living organism using PCR.
2) In vivo cloning - where the gene copies are made within a living organism. As the organism grows and divides, it replicates its DNA, creating multiple copies of the gene.

Front 50

In Vivo Cloning Stage 1 - The Gene is Inserted into a Vector:

Back 50

1) The DNA fragment is inserted into vector DNA.
2) The vector DNA is cut open using the same restriction endonuclease that was used to isolate the DNA fragment containing the target gene, so the sticky ends of the vector are complementary to the sticky ends of the DNA fragment containing the gene.
3) The vector DNA and DNA fragment are mixed together with DNA ligase. DNA ligase joins the sticky ends of the DNA fragment to the sticky ends of the vector DNA. This process is called ligation.
4) The new combination of bases in the DNA (vector DNA + DNA fragment) is called recombinant DNA.

Front 51

In Vivo Cloning Stage 2 - The Vector Transfers the Gene into Host Cells:

Back 51

1) The vector with the recombinant DNA is used to transfer the gene into cells (host cells).
2) If a plasmid vector is used, host cells have to be persuaded to take in the plasmid vector and its DNA
3) With a bacteriophage vector, the bacteriophage will infect the host bacterium by injecting its DNA into it. The phage DNA then integrates into the bacterial DNA
4) Host cells that take up the vectors containing the gene of interest are said to be transformed.

Front 52

In Vivo Cloning Stage 3 - Identifying Transformed Host Cells:

Back 52

1) Marker genes can be inserted into vectors at the same time as the gene to be cloned, so transformed host cells with contain the gene and the marker gene.
2) Host cells are grown on agar plates, and each cell divides and replicates its DNA, creating small colonies
3) Transformed cells will produce colonies where all cells have the gene and the marker gene.
4) The marker gene can code for antibiotic resistance, or fluorescence.
5) Identified transformed cells are allowed to grow more, producing many copies of the cloned gene.

Front 53

In Vivo Cloning Advantages:

Back 53

1) Cloning in vivo can produce mRNA and protein as well as DNA because it's done in a living cell.
2) Cloning in vivo can also produce modified DNA, modified mRNA or modified protein.
3) Large fragments of DNA can be cloned using in vivo cloning, e.g. between 20 to 45 kilobases of DNA can be inserted into some plasmids and bacteriophages.
4) In vivo cloning can be a relatively cheap method, depending on how much DNA you want to produce.

Front 54

In Vivo Cloning Disadvantages:

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1) The DNA fragment has to be isolated from other cell components.
2) You may not want modified DNA.
3) It can be quite a slow process.

Front 55

In Vitro Cloning (PCR) Advantages:

Back 55

1) In vitro cloning can be used to produce lots of DNA (but not mRNA or protein).
2) The DNA produced isn't modified.
3) This technique only replicates the DNA fragment of interest. This means you don't have to isolate the DNA fragment from host DNA or cell compounds.
4) In vitro cloning is a fast process - PCR can clone millions of copies of DNA in just a few hours.

Front 56

In Vitro Cloning (PCR) Disadvantages:

Back 56

1) It can only replicate a small DNA fragment.
2) You may want a modified product.
3) mRNA and protein aren't made as well.
4) It can be expensive if you want to produce a lot of DNA.

Front 57

Genetic Engineering:

Back 57

1) Genetic engineering is the manipulation of an organism's DNA and is also knows as recombinant DNA technology.
2) Organisms that have had their DNA altered are called transformed organisms, with DNA formed by joining together DNA from different sources.
3) Microorganisms, plants and animals can all be genetically engineered to benefit humans.

Front 58

Genetically Engineered Microorganisms:

Back 58

1) Transformed microorganisms can be made using the same technology as in vivo cloning. For example, foreign DNA can be inserted into microorganisms to produce lots of useful protein, e.g. insulin.

Front 59

Genetically Engineered Plants:

Back 59

1) Transformed plants can also be produced - a gene that codes for a desirable characteristic is inserted into a plasmid. The plasmid is added to a bacterium and the bacterium is used as a vector to get the gene into the plant cells. The transformed plant will have the desirable characteristic coded for by that gene.

Front 60

Genetically Engineered Animals:

Back 60

1) Transformed animals can be produced too - a gene that codes for a desirable characteristic is inserted into an animal embryo. The transformed animal will have the desirable characteristic coded for by that gene.

Front 61

Genetic Engineering on Agriculture:

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1) Agricultural crops can be transformed so that they give higher yields or are more nutritious. This means these plants can be used to reduce the risk of famine and malnutrition. Crops can also be transformed to have pest resistance, so that fewer pesticides are needed. This reduces costs and reduces any environmental problems associated with using pesticides.

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Genetic Engineering on Industry:

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1) Industrial processes often use biological catalysts. These enzymes can be produced from transformed organisms, so they can be produced in large quantities for less money, reducing costs.

Front 63

Genetic Engineering on Medicine:

Back 63

1) Many drugs and vaccines are produced by transformed organisms, using recombinant DNA technology. They can be made quickly, cheaply and in large quantities using this method.

Front 64

Issues of Genetic Engineering on Agriculture:

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1) Farmers might plant only one type of transformed crop (monoculture). This could make the whole crop vulnerable to disease because the plants are genetically identical.
2) Some people are concerned about the possibility of 'superweeds' - weeds that are resistant to herbicides. These could occur if transformed crops interbreed with wild plants.

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Issues of Genetic Engineering on Industry:

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1) Without proper labelling, some people think they won't have a choice about whether to consume food made using genetically engineered organisms.
2) Some people are worried that the process used to purify proteins (from genetically engineered organisms) could lead to the introduction of toxins into the food industry.

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Issues of Genetic Engineering on Medicine:

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1) Companies who own genetic engineering technologies may limit the use of technologies that could be saving lives.
2) Some people worry this technology could be used unethically, e.g. to make designer babies. Eugenics are currently illegal though.

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Genetic Engineering Benefits:

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1) Agricultural crops could be produced that help reduce the risk of famine and malnutrition, e.g. drought-resistant crops.
2) Transformed crops could be used to produce useful pharmaceutical products (e.g. vaccines) which could make drugs available to more people.
3) Medicines could be produced more cheaply, so more people can afford them.

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Genetic Engineering Concerns:

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1) Environmentalists - Many oppose recombinant DNA technology because they think it could potentially damage the environment, e.g. monoculture decreasing biodiversity.
2) Anti-Globalisation Activists - A few, large biotechnology companies control some forms of genetic engineering. As the use of this technology increases, these companies get bigger and more powerful. This may force smaller companies out of business.

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Genomes:

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1) Not all of an organism's genome codes for proteins.
2) Some of the genome consists of repetitive, non-coding base sequences. The number of times these sequences are repeated differs from person to person
3) The repeated sequences occur in lots of places in the genome. This can be compared between individuals - genetic fingerprinting.
4) The probability of two individuals having the same genetic fingerprint is very low.

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Electrophoresis Separates DNA Fragments to Make a Genetic Fingerprint:

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1) A sample of DNA is obtained, and the PCR is used to make many copies of the areas of DNA that contain the repeated sequences. You end up with DNA fragments where the length corresponds to the number of repeats the person has at each specific position.
2) A fluorescent tag is added to all the DNA fragments so they can be viewed under UV light.

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Process of Electrophoresis:

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1) The DNA mixture is placed into a well in a slab of gel and covered in a buffer solution that conducts electricity.
2) An electrical current is passed through the gel - DNA fragments are negatively charged, so they move towards the positive electrode at the far end of the gel
3) Small DNA fragments move faster and travel further through the gel, so the DNA fragments separate according to size.

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Genetic Fingerprinting - Post-Electrophoresis:

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1) The DNA fragments are viewed as bands under UV light - this is the genetic fingerprint.
2) Two genetic fingerprints can be compared - e.g. if both fingerprints have a band at the same location on the gel it means they have the same number of nucleotides and so the same number of sequence repeats at that place - its a match.

Front 73

Genetic Fingerprinting - Determining Genetic Relationships:

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1) We inherit the repetitive, non-coding base sequences from our parents. Roughly half of the sequences come from each parent. This means the more bands on a genetic fingerprint that match, the more close related two people are. E.g. paternity tests.

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Genetic Fingerprinting - Determining Genetic Variability Within a Population:

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1) The greater the number of bands that don't match on a genetic fingerprint, the more genetically different people are. This means you can compare the number of repeats at several places in the genome for a population to find out how genetically varied that population is.

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Genetic Fingerprinting - Forensic Science:

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1) The DNA is isolated from all the collected samples.
2) Each sample is replicated using PCR, and the products are run on an electrophoresis gel and the fingerprints produced are compared to see if any match.
3) If the samples match, it links a person to the crime scene.

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Genetic Fingerprinting - Medical Diagnosis:

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1) In medical diagnosis, a genetic fingerprint can refer to a unique pattern of several alleles.
2) It can be used to diagnose genetic disorders and cancer. It's useful when the specific mutation isn't known or where several mutations could have caused the disorder, because it identifies a broader, altered genetic pattern.

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Genetic Fingerprinting - Animal and Plant Breeding:

Back 77

1) Genetic fingerprinting can be used on animals and plants to prevent inbreeding which causes health, productivity and reproductive problems. Inbreeding decreases the gene pool which can lead to an increased risk of genetic disorders, leading to health problems etc.
2) Genetic fingerprinting can be used to identify how closely-related individuals are, so the least related individuals will be bred together.

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Looking For Genes - DNA Probes and Hybridisation:

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1) DNA probes can be used to locate genes or see if a person's DNA contains a mutated gene.
2) DNA probes are short strands of DNA. They have a specific base sequence that's complementary to the base sequence of part of a target gene. This means a DNA probe will bind (hybridise) to the target gene if it's present in a sample of DNA.
3) A DNA probe also has a label attached, so that it can be detected - radioactive or fluorescent.

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Process of DNA Probing and Hybridisation:

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1) A sample of DNA is digested into fragments using restriction endonucleases and separated using electrophoresis.
2) The fragments are then transferred to a nylon membrane and incubated with the fluorescently labelled DNA probe. If the gene is present, the DNA probe will hybridise to it.
3) The membrane is then exposed to UV light and if the gene is present there will be a fluorescent band.

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Restriction Mapping:

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1) Different restriction enzymes are used to cut labelled DNA into fragments.
2) The DNA fragments are then separated by electrophoresis.
3) The size of the fragments produced is used to determine the relative locations of cut sites.
4) A restriction map of the original DNA is made - a diagram of the piece of DNA showing the different cut sites, and so where the recognition sites of the restriction enzymes used are found.

Front 81

Gene Sequencing:

Back 81

1) Gene sequencing is used to determine the order of bases in a section of DNA. It can be carried out by the chain termination method, which lets you sequence small fragments of DNA, up to 750 base pairs.

Front 82

Gene Sequencing - Stage 1:

Back 82

The following mixture is added to four separate tubes:
1) A single-stranded DNA template - the DNA to be sequenced.
2) DNA polymerase - the enzyme that joins DNA nucleotides together.
3) Lots of DNA primer - short pieces of DNA.
4) Free nucleotides
5) Fluorescently-labelled modified nucleotide - like a regular nucleotide, but once it's added to a DNA strand, no more bases can be added after it. A different modified nucleotide is added to each tube (A*, T*, C*, G*)

Front 83

Gene Sequencing - Stage 2:

Back 83

1) The tubes undergo PCR, which produces many strands of DNA. The strands are different lengths because each one terminates at a different point depending on where the modified nucleotide was added.
2) The DNA fragments in each tube are separated by electrophoresis and visualised under UV light.
3) The complementary base sequence can be read from the gel. The smallest nucleotide is at the bottom of the gel. Each band after this represents one more base added. So by reading the bands from bottom to top, you can build up the DNA sequence.

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Sickle-Cell Anaemia:

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1) This is a recessive genetic disorder caused by a mutation in the haemoglobin gene.
2) The mutation causes an altered haemoglobin protein, which makes red blood cells sickle-shaped. These block capillaries and restrict blood flow.
3) Some people are carriers. Sickle-cell carriers are partially protected from malaria. This advantageous effect has caused an increase in frequency of the allele in areas where malaria is common. However, this also increases the likelihood of people inheriting two copies, meaning more people will suffer from sickle-cell anaemia in these areas.

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DNA Probes - Screening for Mutated Genes:

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DNA probes can be used to screen for clinically important genes. There are two ways to do this:
1) The probe can be labelled and used to look for a single gene in a sample of DNA.
2) Or the probe can be used as part of a DNA microarray, which can screen lots of genes at the same time.

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DNA Microarray:

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1) A DNA microarray is a glass slide with microscopic spots of different DNA probes attached to it in rows.
2) A sample of labelled human DNA is washed over the array, and if the labelled human DNA contains any DNA sequences that match any of the probes, it will stick to the array. The array is then washed to remove any labelled DNA that hasn't stuck to it.
3) The array is then visualised under UV light - any labelled DNA attached to a probe will fluoresce.
4) Any spot that fluoresces means that the person's DNA contains that specific gene.

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Genetic Counselling:

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1) Genetic counselling is advising patients and their relatives about the risks of genetic disorders.
2) It involves advising people about screening and explaining the results of a screening. Screening can help to identify the carrier of a gene, the type of mutated gene they're carrying and the most effective treatment.
3) If the results of a screening are positive, then genetic counselling is used to advise the patient on the options of prevention or treatment available.

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Gene Therapy:

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1) Gene therapy involves altering the defective genes inside cells to treat genetic disorders and cancer.
2) If it's caused by two mutated recessive alleles you can adda working dominant allele, or if it's caused by a mutated dominant allele you can 'silence' the dominant allele.
3) The allele is inserted into cells using vectors, and different vectors can be used, e.g. altered viruses, plasmids or liposomes.

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Somatic Therapy:

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1) This involves altering the alleles in body cells, particularly the cells that are most affected by the disorder.
2) For example, cystic fibrosis (CF) is a genetic disorder that's very damaging to the respiratory system, so somatic therapy for CF targets the epithelial cells lining the lungs.
3) Somatic therapy doesn't affect the individual's sex cells though, so any offspring could still inherit the disease.

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Germ Line Therapy:

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1) This involves altering the alleles in the sex cells.
2) This means that every cell of any offspring produced from these cells will be affected by the gene therapy and they won't suffer from the disease.
3) Germ line therapy in humans is currently illegal though.

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Advantages of Gene Therapy:

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1) It could prolong the lives of people with genetic disorders and cancer.
2) It could give people with genetic disorders and cancer a better quality of life.
3) Carriers of genetic disorders may be able to reproduce without passing on the disorder.
4) It could decrease the number of people that suffer from genetic disorders and cancer.

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Disadvantages of Gene Therapy:

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1) The effects of the treatment may be short-lived.
2) The patients might have to undergo multiple treatments.
3) It might be difficult to get the allele into specific body cells, and the body could identify vectors as foreign bodies and initiate an immune response.
4) An allele could be inserted into the wrong place in the DNA, possibly causing more problems.
5) An inserted allele could get overexpressed, producing too much of the missing protein.
6) Disorders caused by multiple genes would be difficult to treat with this technique.

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Ethical Issues with Gene Therapy:

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1) Some people are worried that the technology could be used in ways other than for medical treatment, such as cosmetic effects of ageing.
2) There's the potential to do more harm than good (e.g. risk of overexpression of genes).