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A-Level Biology > 21 Manipulating Genomes > Flashcards

21 Manipulating Genomes Flashcards

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1
Q

What is a genome?

A

The genome of an organism is all of the genetic material it contains.

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2
Q

What are the techniques for studying genes?

A
  • The polymerase chain reaction
  • Cutting out DNA fragments using restriction enzymes
  • Gel electrophoresis
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3
Q

What is PCR used for?

A

PCR is used to select a fragment of DNA and amplify it to copies.
- E.g in crime scene when only tiny amounts of DNA may be available.

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4
Q

What is placed in the PCR machine?

A
  • DNA sample to be amplified
  • Excess of 4 nucleotides
  • Small primer DNA sequences
  • Enzyme DNA polymerase
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5
Q

What happens in Step 1 of PCR?

A

STRANDS SEPERATED

  • The temperature in the PCR machine is increased to 95°C for 30 seconds.
  • This denatures the DNA by breaking the hydrogen bonds holding the DNA strands together so they separate.
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6
Q

What happens in Step 2 of PCR?

A

ANNEALING OF THE PRIMERS

  • The temperature is decreased to 55°C and the primers bind (anneal) to the ends of DNA strands.
  • They are needed for the replication of the strands to occur.
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7
Q

What happens in Step 3 of PCR?

A

SYNTHESIS OF DNA

  • The temperature is increased to 72°C for at least one minute, this is the optimum temperature for DNA polymerase to work best.
  • DNA polymerase adds bases to the primer, building up complementary strands of DNA and so producing double-stranded DNA identical to the original sequence.
  • The enzyme Taq polymerase is used.
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8
Q

As well as PCR, what is another way to get a DNA fragment?

A

Via restriction enzymes.

  • Some sections of DNA have palindromic sequences of nucleotides.
  • These sequences consist of antiparallel base pairs (read in opposite directions)
  • Restriction enzymes recognise specific palindromic sequences and cut the DNA at these places.
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9
Q

What are sticky ends?

A

Small tails of unpaired bases at each end of the fragment.

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10
Q

Why are sticky ends useful?

A

Sticky ends can be used to anneal the DNA fragment to another piece of DNA that has sticky ends with complementary sequences.

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11
Q

What is electrophoresis?

A
  • Electrophoresis is a technique that uses an electrical current to separate out DNA fragments.
  • DNA fragments are put into wells in agarose gel strips which also contain a buffering solution to maintain constant pH.
  • The gel is then placed in an alkaline solution to denature the DNA fragments.
  • The two DNA strands of each fragment seperate.
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12
Q

Why is electrophoresis used?

A

Electrophoresis forms the basis of DNA profiling- a process that can be used to determine how closely related two or more organisms are.

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13
Q

What is DNA profiling?

A

DNA profiling produces an image of the patterns in non-coding DNA of an individual.

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14
Q

What are the uses of DNA profiling?

A

1) FORENSIC SCIENCE: Forensic scientists use DNA profiling to compare samples of DNA collected from crime scenes to samples of DNA from possible suspects, to link them from crime scenes.
2) MEDICAL DIAGNOSIS: Can be used to analyse risk of genetic disorders.

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15
Q

How is a DNA profile produced?

A

1) Extracting DNA- DNA extracted from tissue sample, or amplified using PCR
2) Digesting sample- Strands of DNA are cut into small fragments using restriction endonucleases. These cut DNA at a recognition side (specific nucleotide sequence)
3) Separating DNA fragments- Cut fragments of DNA are separated using gel electrophoresis. DNA fragments are transferred from the gel to nylon membrane via Southern blotting.
4) Hybridisation- Radioactive or fluorescent DNA probes are added in excess to DNA fragments on the membrane. DNA probes are short DNA sequences complementary to the sequences.
5) If Radioactive labels added- X-ray images taken
If fluorescent labels added- Membrane placed under UV light so fluorescent tags show.
The fragments give a pattern of bars- DNA profile.

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16
Q

What is computational biology?

A

The study of biology using computational techniques to analyse large amounts of data.

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17
Q

What is bioinformatics?

A
  • The development of software and computing tools needed to analyse and organise raw biological data.
  • Computational biology then uses this data to build theoretical models of biological systems which predicts what will happen in different circumstances.
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18
Q

What is genomics?

A

The field of genetics that applies DNA sequencing methods and computational biology to analyse the structure and function of genomes.

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19
Q

What are the uses of genome sequencing?

A
  • Analysing the human genome
  • Analysing the genomes of pathogens
  • Identifying species
  • Searching for evolutionary relationships
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20
Q

What are the benefits of analysing the human genome?

A
  • Computers can analyse and compare the genomes of many individuals, revealing patterns in the DNA we inherit and the diseases to which we are vulnerable.
21
Q

What are benefits of analysing the genomes of pathogens?

A
  • Can find out the source of an infection
  • Can identify antibiotic-resistant strains of bacteria and plan suitable treatment options.
  • Can track the progress of an outbreak of a potentially serious disease and monitor potential epidemics
  • Can identify regions in the genome of pathogens that may be useful targets in the development of new drugs and to identify genetic markers for use in vaccines.
22
Q

What is DNA barcoding?

A
  • When scientists identify species using relatively short sections of DNA from a conserved region of the genome.
  • This section is small enough to be sequenced quickly and cheaply, yet varies enough to give clear differences between species.
23
Q

What is proteomics?

A

Proteomics is the study and amino acid sequencing of an organisms entire protein complement.

24
Q

What are spliceosomes?

A
  • Spliceosomes are enzyme complexes that join exons together to give the mature functional mRNA.
  • The spliceosomes may join the same exons in a variety of ways.
  • As a result, a single gene may produce several versions of functional mRNA, which in turn codes for different arrangements of amino acids, giving different proteins, resulting in several different phenotypes.
25
Q

What is synthetic biology?

A

The design and construction of novel biological pathways, organisms or devices, or the redesign of existing natural biological systems.

26
Q

What are the techniques of synthetic biology?

A
  • Genetic engineering
  • Use of biological systems in industrial contexts e.g production of drugs from microorganisms
  • Synthesis of new genes to replace faulty genes
  • Synthesis of a new organism
27
Q

How is DNA sequencing carried out?

A

1) The DNA for sequencing is mixed with a primer, DNA polymerase, an excess of normal nucleotides (containing bases A, T, C and G) and terminator bases.
2) The mixture is placed in a thermal cycler, and at 96°C the double stranded DNA separates into single strands, at 50°C the primers anneal to the DNA strand.
3) At 60°C DNA polymerase starts to build up new DNA strands by adding nucleotides with the complementary base to the single strand DNA template.
4) Each time a terminator base is incorporated instead of a normal nucleotide, the synthesis of DNA is terminated as no more bases can be added.
- As the chain-terminating bases are present in lower amounts and are added at random, this results in many DNA fragments of different lengths depending on where the chain terminating bases have been added during the process.
- After many cycles, all of the possible DNA chains will be produced with the reaction stopped at every base.
- The DNA fragments are separated according to their length by capillary sequencing, which works like gel electrophoresis in minute capillary tubing.
- The fluorescent markers on the terminator bases are used to identify the final base on each fragment.
- Lasers detect the different colours and thus the orders of the sequence.
5) The order of bases in the capillary tubes shows the sequence of the new, complementary strand of DNA which has been made.
- This is used to build up the sequence of the original DNA strand.
- The data from the sequencing process is fed into a computer that reassembles the genomes by comparing all the fragments and finding the areas of overlap between them.
- Once a genome is assembled, scientists identify the genes that code for specific characteristics.

28
Q

What is next generation sequencing?

A

When all of the clusters are being sequenced and imaged at the same time.

29
Q

What are the basic principles of genetic engineering?

A

Isolating a gene for a desirable characteristic in one organism and placing it into another organism, using a suitable vector.

30
Q

What does transgenic/genetically modified organism mean?

A

An organism that carries a gene from another organism

31
Q

What is the first stage of genetic engineering?

A

Isolating the desired gene.

  • Enzymes called restriction endonucleases cut the required gene from the DNA of an organism. It cuts the two DNA strands unevenly producing sticky ends. The sticky ends make it much easier to insert the desired gene into the DNA of a different organism.
  • Another technique involves isolating the mRNA for the desired gene and using the enzyme reverse transcriptase to convert mRNA go gene to cDNA (more time efficient as introns have been removed, already modified)
32
Q

What is the most commonly used vector in genetic engineering?

A

Plasmids- small circular molecules of DNA separate from the chromosomal DNA that can replicate independently.

33
Q

What occurs once a plasmid gets into a new host cell?

A

It combines with the host DNA to form recombinant DNA.

34
Q

How is a DNA fragment inserted into a plasmid?

A

1) The same restriction endonuclease used to isolate the DNA fragment is used to cut the plasmid.
2) This results in the plasmid having complementary sticky ends to the sticky ends of the DNA fragment.
3) Once the complementary bases of the two sticky ends are lined up, the enzyme DNA ligase forms phosphodiester bonds between the sugar and phosphate groups on the two strands of DNA, joining them together.
4) The plasmids used as vectors are usually given a second marker gene, which is used to show that the plasmid contains the recombinant gene.

35
Q

How is the plasmid with the recombinant DNA transferred into the host cell?

A

There are two ways:

1) The bacterial cells and plasmids are cultured in a calcium rich solution and the temperature is increased. This causes the bacterial membrane to become permeable and the plasmids can enter.
2) Electroporation: The use of a very tiny electric current to transfer genetically engineered plasmids into bacteria by making the membrane very porous. It is also used to get DNA fragments directly into eukaryotic cells.

36
Q

What is another way of producing genetically modified cells?

A

Electrofusion.

  • In electrofusion, tiny electric currents are applied to the membranes of two different cells.
  • This fuses the cell and nuclear membranes of the two different cells together to form a hybrid or polyploid cell containing DNA from both.
37
Q

How is insect resistance brought about in soya beans?

A
  • Scientists inserted a gene into soya beans so they produce the Bt protein.
  • This protein is toxic to many pest insects that attack the plant.
38
Q

What are the pros of pest resistance in GM crops?

A
  • Reduce the amount of pesticide spraying, protecting the environment and helping poor farmers.
  • Increased yield
39
Q

What are the cons of pest resistance in GM crops?

A
  • Non-pest insects might be damaged by the toxins in the GM plants.
  • Insect pests may become resistant to the pesticides in GM crops.
40
Q

What are the pros of disease resistance in GM crops?

A
  • Reduces crop losses and increases yield
41
Q

What are the cons of disease resistance in GM crops?

A
  • Transferred genes might spread to wild populations and cause issues e.g superweeds
42
Q

What are the pros of herbicide resistance in GM crops?

A
  • Herbicides can be used to reduce competing weeds and increase yields.
43
Q

What are the cons of herbicide resistance in GM crops?

A
  • Biodiversity could be reduced if herbicides are overused to destroy weeds.
44
Q

What is the issue with patenting and technology transfer?

A
  • When someone discovers a new technique and legally patents it, it means no one else can use it without payment.
  • The people who need benefits of drought/resistant crops and high yields the most might be unable to afford the GM seed, especially if they’re in a less economically developed country.
45
Q

What is pharming?

A
  • The use of genetically modified animals to produce pharmaceuticals.
46
Q

What are two aspects of pharming?

A
  • Creating animal models: adding or removing genes so that animals develop certain diseases, acting as models for development of new therapies
  • Creating human proteins- animals are used to produce milk containing human proteins that can be harvested.
47
Q

What are the ethical issues with researching microorganisms?

A
  • Some people are worried that scientists researching the pathogen could become infected with the live pathogen and potentially cause a mass outbreak of disease.
  • The genetically modified version of a pathogen could revert back to its original form and cause an outbreak of disease.
48
Q

What is somatic cell gene therapy?

A

Replacing a faulty gene with a healthy allele in affected somatic cells.

49
Q

What is germ line cell gene therapy?

A

Inserting a healthy allele into the germ cells or into a very early embryo.

A-Level Biology (41 decks)

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