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6.1.3 Manipulating genomes

Zoe Mitchell Biology A level English 94 cards

Cards in this deck (94)

  1. 1DNA sequencinga technique that allows genes to be isolated and read
  2. 2Who was the british biochemist that developed what technique that led to advancements which allowed scientist to sequence whole genomes?Fred Sanger
    Sanger's chain termination DNA sequencing
  3. 3How do you set up Sanger's DNA sequencing approach?Put a single-stranded DNA template, DNA primer, DNA polymerase and free nucleotides into 4 tubes. Into each of the tubes, put one of type of DNA radioactive modified nucleotide. Once synthesised into the complementary DNA, no more bases can be added to the DNA strand.
  4. 4Explain how Sanger's DNA sequencing approach works.Primers anneal to the 3' end and DNA polymerase attaches. Polymerase chain reaction occurs. When a modified nucleotide attaches, no more bases can attach, producing many diffrent length strands of DNA. The base at the end of each strand is known because it is known in which tube the DNA was synthesised. Separated by electrophoresis and the DNA is built up base by base.
  5. 5What was the first DNA sequencing machine?based on Sanger's method but nucleotides were labelled with fluorescent dyes, instead of radioactive labels. A laser beam scanned the dyes and the light signature was recognised by the computer.
  6. 6DNA sequencing by the chain termination method only works on fragments up to ? base pairs long.750
  7. 7A whole genome is too long to be sequenced by chain termination method. How could a whole genome be sequenced still using this method?Cut the genome into shorter fragments using restriction enzymes. insert into man-made plasmids and then inserted into bacteria which create clonanies of clones. Extract DNA from bacteria. Cut up using restriction enzymes. Sequence using chain-termination method. Computer puts DNA fragments back into order. Whole genome has been sequenced
  8. 8Man-made plasmids akabacterial artificial chromosomes
  9. 9high-throughput sequencingtechniques that can sequence DNA much faster and much more cheaply compared to Sanger's chain termination method. For instance, pyrosequencing.
  10. 10Explain how pyrosequencing works.DNA cut into fragments by a nebuliser. Split into single-stranded DNA & immobilised. Sequencing primer, DNA polymerase and enzymes which cause light to be emitted when a nucleotide is added to the DNA strand and apyrase are all added to the single stranded DNA. One of the four possible nucleotides with two extra phosphoryl groups is added one at a time. When one of the nucleotides is added to the DNA strand, the phosphoryl groups are converted to light by special enzymes. The light is detected and the amount of light is proportional to how many of the same type of nucleotide are adjacent. Unincorporated nucleotides are degraded by apyrase and the reaction is repeated with another nucleotide.
  11. 11What reagents are used in pyrosequencing?single stranded DNA fragments
    sequencing primer
    DNA polymerase
    enzymes which cause light to be emitted when a nucleotide is added to the DNA strand
    aparase
  12. 12How are DNA nucleotides modified for use in pyrosequencing?The nucleotides have 2 extra phosphoryl groups
  13. 13How are the DNA nucleotides added in pyrosequencing?Only one type of nucleotide is added at a time.
  14. 14What happens in pyrosequencing if a nucleotide is added but is not complementary to the template DNA so is not used?Unincorporated nucleotides are degraded by apyrase and the reaction is repeated with another nucleotide.
  15. 15What happens in pyrosequencing when nucleotides are added?When one of the nucleotides is added to the DNA strand, the phosphoryl groups are converted to light by special enzymes. The light is detected and the amount of light is proportional to how many of the same type of nucleotide are adjacent.
  16. 16How long is a normal run in pyrosequencing? How many bases can pyrosequencing sequence in a normal run?10-hour run
    400 million bases
  17. 17Bioinformaticsdeveloping and using computer software that can analyse, organise and store biological data. It has allowed the huge amounts of data generated from genome sequences to be stored.
  18. 18Computational biologyusing computers to study biology such as creating computer simulations and mathematical models
  19. 19Uses of gene sequencingComparisons between individuals & species
    Predicting the primary structure of polypeptides
    Synthetic biology
  20. 203 uses of comparisons between indivduals & species using gene sequencingGenotype-phenotype relationships
    Epidemiology
    Evolutionary relationships
  21. 21How has gene sequencing helped investigate genotype-phenotype relationships?by analysing an organism’s genotype, their phenotype can be predicted. Useful in the diagnosis and treatment of genetic disorders.
  22. 22What is epidemiology and why is it useful?the study and analysis of the patterns, causes, and effects of health and disease conditions in a defined population. Uses genome sequencing to understand why genetically similar people suffer from different diseases which can help detect the mutations which cause or increase the risk of a disease
  23. 23How can gene sequencing be used to improve the understanding of evolutionary relationships?comparing genomes of different species helps to show how closely related they are and when the species evolved away from each other. Comparing genomes of the same species can show when different groups of the organism separated and how their genomes changed in slightly different ways.
  24. 24How can gene sequencing be used to figure out the primary structure of polypeptides?knowing the sequence of bases in a gene means that the sequence of amino acids coded for by triplets of bases can be found out
  25. 25Synthetic biologyinterdisciplinary science concerned with designing and building useful biological molecules, devices and systems from scratch.
  26. 264 examples of synthetic biology-building artificial molecules to test how they work i.e. proteins
    -producing medicines
    -redesigning systems to make them work better
    -designing new useful biological systems & molecules that don't excist naturally
  27. 27Give an example of a medicine that synthetic biology has madeartemisinin – an antimalarial drug – originally produced in plants, is now produced by a genetically engineered precursor – yeast.
  28. 28Give an example of a redesigned biology system that synthetic biology has producedredesigning haemoglobin so that it binds to oxygen, but not to carbon monoxide
  29. 29Give an example of a new biological molecule/system that doesn't exsist naturally which synthetic biology may produce.energy products to use as fuel
  30. 30Synthetic biology is reliant on _____ & _____ to ____bioinformatics & computational biology
    create computational simulations, mathematical models, store & analyse data
  31. 31tandem repeatsrepetitive segments of DNA which do not code for proteins
  32. 32Tandem repeats are unique in each individual because ....the number of times a sequence is repeated at more than 1000 gene loci, each for a different, random number of repeats
  33. 33What is a DNA profile?When DNA has undergone electrophoresis to analyse how many times the tandem repeats are repeated in a genome at certain loci.
  34. 34How is DNA profiling carried out?1. DNA obtained from individual
    2. Restriction enzymes used to cut the DNA at specific recognition sites into varying length fragments depending on the individual
    3. Fragments are separated by gel electrophoresis and stained
    4. Banding pattern is compared to other genome’s treated to the same restriction enzymes and gel electrophoresis
    5. If the bands match, they are from the same genome
  35. 35name 5 sources of DNA which can be used for DNA profilingsaliva, blood, hair, skin cells, semen
  36. 36How is DNA profiling used in forensic science?compare samples of DNA from a crime scene to the DNA from suspects
  37. 37How is DNA profiling used in analysis of disease risk?Used to aid disease risk when the specific mutation for a disease isn’t known or if several mutations could cause the disease i.e. Hungtington's diseases, sickle cell anaemia
    Also used in IVF to screen an embryo's DNA to compare to the faulty regions of the parent's DNA i.e. Huntington's disease, cystic fibrosis
  38. 38What is PCR?Polymerase Chain Reaction – reaction used to artificially replicate many copies of a DNA fragment – amplification
  39. 39features of DNA that allow PCR to work2 antiparallel backbone strands
    Each strand has a 5’ and a 3’ end
    DNA extends from the 3’ only
    Complementary base pairing rules
  40. 40What are primers?short single strand of DNA that are complementary to the bases at the start of the fragment you want
  41. 41What is Taq DNA polymerase?enzyme from a bacteria which catalyses the addition of DNA nucleotides to the single-stranded DNA which is stable at high temperatures
  42. 42annealbind by hydrogen bonds
  43. 43What does the initial reaction mixture of a polymerase chain reaction contain?DNA sample, free nucleotides, primers, Taq DNA polymerase
  44. 44Explain the process of PCR.Reaction mixture heated to 95 C to break hydrogen bonds between DNA strands making 2 single strands of DNA. Cooled to 55 C Primers anneal to one end of the single strands. Heat to 72C - optimum temp for Taq polymerase which catalyses addition of free nucleotides to the single strand DNA, starting by the primer, moving in the 5' to 3' direction. Double stranded DNA made. Cycle repeated.
  45. 45How does the amount of DNA in PCR increase?exponentially
  46. 465 Differences between PCR and natural DNA replicationIn PCR:
    -Only short strands, not whole chromosomes, replicated
    -Requires primers because DNA is completely broken into single strands
    -Heat, not helicase, used to separate DNA strands
    -Taq DNA polymerase, not DNA polymerase, used
    -Requires cycle of heating and cooling to precise temps, rather than at temp of the organism
  47. 47application of PCRreplicating DNA for forensic science, detecting mutations, research into extinct ancient DNA sources
  48. 48electrophoresisprocess used to separate proteins or DNA fragments of different sizes
  49. 49What gel is used in electrophoresis?agarose gel
  50. 50Describe how you set up the gel box for electrophoresis.1. Pour agarose gel into a gel tray with a comb placed at one end. Leave to solidify. 2. Remove the comb from the gel, creating a row of wells. 3. Put the gel tray into a gel box, making sure the wells are at the negative electrode. 4. Add a buffer solution to the gel box, ensuring the gel tray is covered and that the reservoirs contain buffer.
  51. 51Why does electrophoresis work?DNA has an overall negative charge. All fragments, regardless of size, have the same charge. Therefore, fragments will be attracted towards the positive electrode.
  52. 52How do you put the DNA samples into the electrophoresis apparatus?Add loading dye to a fragmented DNA sample. Add a set volume of the DNA sample to the wells using a micropipette. Make sure it doesn’t pierce the bottom of the wells. Using a clean micropipette tip each time, add the other DNA samples to the other wells.
  53. 53What do you do in electrophoresis after you have loaded the DNA sample?Put the lid on the gel box and connect the electrodes to the power supply. Set the voltage of the power supply and turn it on. Leave until the dye is about 2cm from the end of the gel. Turn off the supply and remove the gel tray. Tip off excess buffer solution. Stain the DNA fragments by covering the gel surface with a staining solution. View the bands of DNA fragments.
  54. 54If electrophoresis is carried out on proteins, instead of DNA, what extra step has to be carried out?mixed with a chemical that denatures the proteins so that they all have the same charge
  55. 55Uses of electrophoresisSeparate proteins to identify proteins in urine or blood to diagnose diseases such as sickle cell anaemia and leukaemia
    DNA profiling for forensic science & maternity & paternity disputes
  56. 56List the 4 main steps of genetic engineering1. The DNA fragment containing the required gene is obtained
    2. A copy of the gene is placed inside a vector
    3. The vector carries the gene into the recipient cell
    4. The transformed cell expresses the desired gene
  57. 57In genetic engineering how is the DNA fragment containing the required gene obtained?Restriction enzymes cut out the required gene. The active site of the enzyme is complementary to the shape of the recognition sequence where it cuts. The cuts leave sticky ends.
  58. 58restriction enzymesendonuclease enzymes that cleave DNA molecules at specific recognition sites
  59. 59sticky endssmall tails of unpaired bases at each end of the fragment which can be used to help bind to DNA with ends which have complementary sequences
  60. 60vectorin gene technology anything that can transfer DNA into a host organism
  61. 612 examples of a vectorplasmids or bacteriophages (viruses which infect bacteria.)
  62. 62Plasmidssmall loops of DNA in prokaryotic cells
  63. 63DNA ligaseenzyme that catalyses the joining of sugar and phosphate groups within DNA
  64. 64Recombinant DNAa composite DNA molecule created in vitro by joining foreign DNA fragment with vector DNA
  65. 65Explain how a copy of the gene is placed inside a vector in genetic engineeringVector DNA is cut open with the same restriction enzymes that were used to isolate the DNA fragment, making complementary sticky ends to the DNA fragment. The DNA fragment, the vector DNA and DNA ligase are mixed together and the DNA fragment and the vector DNA anneal, catalysed by DNA ligase, making recombinant DNA.
  66. 66Explain how a plasmid vector carries the gene into the recipient cellElectroporation - A high voltage pulse applied to the cell makes the recipient cell membrane more porous so the plasmids can enter the cell.
  67. 67Explain how a plasmid bacteriophage carries the gene into the recipient cellit will infect the bacterium by inserting its genome into the host cell genomes.
  68. 68transgenicorganism containing a gene from another species
  69. 694 ethical issues to do with genetic modification-insect resistance in GM soya
    -GM pathogens for research
    -GM animals for pharming
    -Patenting
  70. 70Explain how GM soya beans are insect resistantgenetically modified to include a gene (originally from a bacteria) that produces a protein that is toxic to insects that feed on the soya bean plants
  71. 71Positive ethical issues about insect resistant soya beansIncrease yield so more food produced
    reduces amount that chemical pesticides are used
  72. 72Negative ethical issuesMay encourage a monoculture, decreasing biodiversity and leaving the crop vulnerable to disease
    Weeds may become resistant to the toxin, producing superweeds
  73. 73Pros of using GM pathogens for researchPrevious untreatable disease may be able to be cured
  74. 74Cons for using GM pathogens for researchMay cause a mass outbreak of disease
    May be used to create agents for biowarfare
  75. 75pharmingusing genetically modified animals to produce pharmaceuticals
  76. 76pros of pharmingLarger quantities may be produced
    Animals are used where the protein needed is too large for bacteria to produce
  77. 77Cons of pharmingAnimals may suffer harmful side-effects
    Concern about how the animals are looked after
  78. 78Patentingcompanies can control who uses the product and how they use it, for a set period of time
  79. 79Pros of patenting in GM p• Owner will generate money from selling product which may fund further research
    • May encourage technology to advance faster as scientist compete to come up with it first
  80. 80Cons of patenting in GMFarmers in poorer countries may not be able to afford patented GM seeds
  81. 81Example of research carried out with GM pathogenssome tumour cells have receptors on their membranes for poliovirus. Poliovirus will recognise and attack these cells. If poliovirus could be inactivated from causing poliomyelitis, it may be able to become a treatment for cancer.
  82. 82Example of pharminghereditary antithrombin deficiency causes blood clots. Treatment is by infusion of the protein antithrombin. Genetically engineered goats can produce high yields of this protein. It is extracted and used to produce a drug given to people with this disease.
  83. 83gene therapytreatment for genetic disorders that works by altering alleles within cells
  84. 84If the genetic disorder is caused by 2 recessive alleles, explain the principle behind gene therapya dominant allele may be able to be put into cells so that these cells are then able to make the missing protein
  85. 85If the genetic disorder is caused by a dominant allele, explain the principle behind gene therapythe dominant allele could be ‘silenced’ by using interference RNA to block translation.
  86. 86What type of vectors may be used in gene therapy?liposomes (small spheres of lipid bilayer which contain the alleles), altered viruses or plasmids.
  87. 872 types of gene therapygerm line therapy
    somatic cell therapy
  88. 88germ line therapygene therapy by inserting functional alleles into gametes or zygotes. Every cell in the individual will be altered, and offspring may inherit the foreign alleles and the disease won’t be passed on to offspring.
  89. 89somatic cell gene therapygene therapy by inserting functional alleles into body cells. Only affect certain cells. Doesn’t affect gametes so faulty genes could be passed on to offspring.
  90. 90Give an example of somatic cell gene therapyIn cystic fibrosis gene therapy, liposomes with the missing dominant alleles are sprayed into the nose. Some will pass through the plasma membrane and nuclear envelope of the cells lining the respiratory tract and insert into the host genome, the cell will express the protein which the cells normally can’t produce, reducing symptoms of CF. Cells are replaced within a few days, so this treatment needs to be repeated.
  91. 912 pros of somatic cell gene therapyCould prolong life of people with genetic disorder
    Could improve quality of life for people with genetic disorders
  92. 922 pros for germ line gene therapysufferers could conceive a baby without the risk of the genetic disorder
    could decrease the number of people with genetic disorders
  93. 93Cons of somatic cell gene therapyresults may be short-lived and have to be repeated regularly
    may be difficult to get the allele into specific body cells
    May initiate an immune response to the vectors
    May cause cancer, overexpression of a certain protein or other illnesses
    Expensive
  94. 94Cons of germ line gene therapyMay cause cancer, overexpression of a certain protein or other illnesses
    Expensive
    May be used for other purposes other than medical treatment, such as cosmetic uses or ‘designer’ babies
    patient can’t give consent because they are not yet alive

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A level Biology (Module 6: Genetics, evolution and ecosystems) Flashcards on 6.1.3 Manipulating genomes, created by Zoe Mitchell on 01/04/2017.
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