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Biology 🧬 > gene technology > Flashcards

gene technology Flashcards

1
Q

what is recombinant DNA technology?

A

transfer of fragments of DNA from one organism to another
DNA that contains nucleotides from other organisms

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2
Q

3 ways of producing DNA fragments

A
  1. reverse transcriptase
  2. restriction endonucleases
  3. gene machine
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3
Q

how can reverse transcriptase make DNA fragments?

A
  1. remove mRNA from a cell
  2. add reverse transcriptase and DNA nucleotides
  3. converts the mRNA into DNA (double stranded)
    uses mRNA as template

= cDNA (complementary)

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4
Q

why is mRNA used for reverse transcriptase?

A

easier to isolate from cells than DNA
introns already removed in splicing

cDNA produced is complementary to mRNA, therefore is a copy of original DNA

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5
Q

how can restriction endonucleases make DNA fragments?

A
  1. obtain sample of DNA from a cell
  2. incubated with specific restriction endonuclease
  3. cuts DNA fragment using hydrolysis
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6
Q

how do restriction endonucleases work?

A
  • different endonucleases cut at different base sequences
  • shape of sequence is complementary to enzymes active site
  • used to obtain gene between the sequences to cut it out, hydrolysis
    (gene cant contain specific sequence or it will be cut)
    leaves sticky or blunt ends
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7
Q

what are sticky and blunt ends?

A

when a gene is cut using a restriction endonuclease, leaves:

sticky ends - one strand of fragment longer than the other
- easier to insert gene as hydrogen bonds can form between bases

blunt ends - when both strands are same length
- nucleotides can be added to create sticky ends

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8
Q

how does a gene machine produce DNA fragments?

A
  1. determine amino acid sequence of the protein
  2. first nucleotide fixed to support
  3. nucleotides added step by step in order
  4. creates oligonucleotides (short sections of DNA)
  5. oligonucleotides joined to make longer fragments
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9
Q

ways of amplifying DNA fragments

A

PCR (in vitro)
in vivo

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10
Q

describe PCR as a method to amplify DNA fragments

A
  1. reaction mixture made
  2. heated to 95c
    - hydrogen bonds break, strands separate
  3. cooled to 55c
    - allows primers to bind
  4. heated to 72c
    - allows complementary nucleotides to attach (optimum)
  5. heat stable (taq) DNA polymerase joins nucleotides together
    - with phosphodiester bonds

repeats
- each PCR cycle doubles amount of DNA

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11
Q

what does the reaction mixture for PCR contain?

A
  • DNA sample
  • free DNA nucleotides
  • primers
  • DNA polymerase (taq)
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12
Q

in vivo method of amplifying DNA fragments

A
  1. use restriction endonuclease to obtain desired gene from DNA
  2. use same RE to cut open a plasmid
    - leaves sticky ends complementary to sticky ends of fragment
  3. join desired gene to plasmid using DNA ligase
    - joins sticky ends

creates recombinant DNA (vector and fragment)

  1. vector (plasmid) inserted into a bacteria cell
  2. plasmid replicated during binary fission to create more recombinant DNA (and therefore gene)
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13
Q

what are transformed cells and how can you identify them?

A

cells that have taken up the vectors, contain plasmids with recombinant DNA

  • marker genes
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14
Q

how do marker genes identify transformed cells?

A

used to identify which cells contain the plasmid with the desired gene

  1. another gene (for an identifiable characteristic) is inserted with the desired gene
  2. means transformed cells also contain that gene
    - eg antibiotic resistance, so only transformed and resistant cells survive

or gene inserted within a gene for antibiotic resistance
tranformed are killed

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15
Q

benefits of recombinant DNA tech in agriculture

A

crops can be transformed to be:
- more nutritious
- higher yields
reduce risk of famine

  • pest resistant (fewer pesticides needed, cheaper)
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16
Q

benefits of recombinant DNA tech in industry

A

industrial process often use enzymes
- can be made from transformed organisms
- made in large quantities and cheaper

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17
Q

benefits of recombinant DNA tech in medicine

A

transformed organisms used to make drugs and vaccines
- can be made cheaper and quicker
eg insulin

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18
Q

issues of recombinant DNA tech in agriculture

A

monoculture of transformed crops - genetically identical
- makes whole crop vulnerable to same disease - famine risk
- reduces biodiversity

long term effects of GM food on health unknown

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19
Q

issues of recombinant DNA tech in industry

A

some consumer markets won’t import genetically modified food - producers suffer economically

no labelling - no choice over eating GM food or not, unethical

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20
Q

issues of recombinant DNA tech in medicine

A

could lead to unethical uses
eg designer babies - modifying and choosing characteristics

can be used to save lives but companies may limit use for economic reasons

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21
Q

humanitarian benefits of recombinant DNA tech

A
  • agricultural crops made that can reduce risk of famine and malnutrition
    eg drought resistant crops
  • used to make vaccines and drugs available to more people
    eg made chpear so more people can afford them
  • can be used in gene therapy to treat diseases
22
Q

environmentalists issues with recombinant DNA tech

A

monocultures (of transformed crops crops) reduce biodiversity

23
Q

anti-globalisation activists issues with recombinant DNA tech

A

oppose globalisation (growth of large multinational companies)

few large companies control forms of genetic engineering
as use increases, these companies get bigger and more powerful
- smaller companies unable to compete

24
Q

what does gene therapy do?

A

used to treat genetic disorders

it alters the defective genes (mutated allele causing disease)

done by inserting DNA fragment into persons original DNA

25
Q

how does gene therapy work to change the mutated gene?

A

if the mutated alleles are:

2 recessive - add dominant to supplement them

1 dominant - silence it (add DNA to middle of allele so it doesn’t work anymore)

26
Q

2 types of gene therapy

A

somatic - altering alles in body cells
particularly those affected by disease
- eg epithelial cells in lungs for cystic fibrosis
(doesn’t affect sex cells, could be inherited)

germ line - altering alleles in sex cells
disease can’t be inherited as every sex cell has been altered

27
Q

how is gene therapy done?

A

working allele inserted into cells using vectors
vectors include:
- altered viruses
- plasmids
- liposomes (spheres of lipids)

28
Q

what is a DNA probe?

A

short strands of DNA
have a specific base sequence complementary to the target allele

means they will bind (hybridise) to the target allele if present

has a label attached so it can be detected
- fluorescence (use UV light)
- radioactive (use x ray film)

29
Q

how does a DNA probe work?

A

DNA fragment obtained
(restriction endonucleases and separated using electrophoresis)
- transferred onto a nylon membrane

sample of DNA incubated with DNA probe (with label)
if target allele present, DNA probe will bind to it
- probe not removed when washed

use UV light or x-ray film to detect DNA probe, if present

DNA in sample must be single stranded to allow the probe to bind

30
Q

what is a DNA probe used for?

A

locating specific alleles of genes

see if persons DNA contains mutated alleles that causes genetic disorders

used for screening

31
Q

uses of screening with DNA probes

A
  1. identify heritable conditions
    screened to identify mutated allele
    - means treatment can be started earlier eg in cystic fibrosis
  2. determine how patients will respond to drugs
    screening for a specific mutated allele can show if someone has it so if a specific drugs will work
  3. identify health risks
    inheriting a particular mutated allele can increase risk of developing certain cancers
    - allow them to make life choices to reduce their risk

however could lead to discrimination by insurance companies or employers if some people have a higher risk of disease

32
Q

uses of DNA probes (3)

A

locate specific alleles
can screen for:
- heritable conditions
- responses to drugs
- health risks

33
Q

what is genetic counselling?

A

advising patients and families about the risk of genetic disorders
advise on screening and the results

  • the risks of it being passed on to offspring
  • advice on future steps eg most effective prevention or treatments
  • explain potential consequences of disease eg economically

screening can help identify if someone is a carrier of a mutated allele and what type (showing what genetic disorder it causes)

34
Q

what is personalised medicine?

A

medicines that are tailored to an individuals DNA

genes mean different people respond to drugs in different ways - makes them more effective for some than others
can be used in prevention if mutated allele detected

if doctors have genetic information it can be used to predict responses to drugs
therefore prescribe the most effective

35
Q

benefits of genetic screening

A

can identify people with allele who are at risk of developing specific diseases
- can make lifestyle choices to reduce their risk eg diet

can help potential parents chose to have children or not
- may not want to risk passing on harmful alleles

36
Q

limitations of genetic screening

A

designer babies - unethical

could lead to discrimination in workplaces or for insurance
- for people who have a higher risk of developing certain diseases

37
Q

why are peoples genetic fingerprints different?

A

genome contains variable number tandem repeats
- base sequences that don’t code for proteins and repeat next to each other

number of times the sequences are repeated = length in nucleotides
- this varies between people

repeats also occur at different places in the genome
- varies between people

probability of 2 people having same genetic fingerprint is very low
- need same number of VNTRs at same place in genome

38
Q

how are the fragments obtained for electrophoresis?

A

restriction endonucleases used to at specific base sequences to cut VNTRs out

results in fragments of varying lengths - depending on number of repeats

apified using PCR = many copies

39
Q

why are the fragments negatively charged?

A

DNA contains a phosphate group
= negative

so attracted to positive electrode at end of gel

40
Q

describe electrophoresis (how a genetic fingerprint is made)

A

sample of DNA obtained (blood, saliva etc)
PCR used to make copies of area of DNA fragment containing VNTRs
electrophoresis used to separate them
fluorescent tag added - visible in UV

  1. DNA mixture placed in well in gel
    covered in a buffer solution that conducts electricity
  2. electric current passed through gel
    DNA fragments negatively charged
    move towards positive electrode at far end of gel
  3. small DNA fragments move faster so travel further
    larger ones don’t travel as far
    - DNA fragments separate according to size

viewed as bands under UV light (fragments had fluorescent tag)

41
Q

How to view a genetic fingerprint

A

Fragments transferred from gel to nylon membrane
Membrane incubated with fluorescently labelled DNA probe
Viewed as bands under UV light

42
Q

what is genetic fingerprinting?

A

method used to determine an organisms genetic identity

done by electrophoresis

depends on number of VNTRs in the organisms DNA
- shown as bands under UV or radioactive light

43
Q

interpretation of a genetic fingerprint

A

multiple fingerprints can be compared

remember fragments separate based on length (in nucleotides) and therefore VNTRs
shorter move further

if both fingerprints have a band at the same location, they have the same number of nucleotides (so same number of VNTRs)
they match

44
Q

uses of genetic fingerprinting in forensic science

A

compare samples of DNA collected from crime scenes to samples from suspects

  1. DNA isolated from crime scene and suspects
  2. each sample replicated using PCR
  3. samples run on electrophoresis gel to produce genetic fingerprints
  4. compared to find a match
    - if samples match, it can link a suspect to a crime scene
45
Q

uses of genetic fingerprinting in medical diagnosis

A

genetic fingerprint refers to a unique pattern of alleles
can be used to diagnose genetic disorders and cancer

useful when specific mutation isn’t known or when caused by multiple mutations
because it identifies a broader genetic pattern

46
Q

uses of genetic fingerprinting in animal and plant breeding

A

used to prevent interbreeding
- decreases gene pool
- increased risk of genetic disorders
- heath and productivity issues

used to identify how closely related two individuals are so they won’t be bred together (less related bred)

more similar fingerprints means more closely related

47
Q

how is genetic fingerprinting used to determine genetic relationships?

A

VNTR sequences inherited from parents
- roughly half from each parent

more bands that match = more closely related

eg paternity tests - more matches = more likely to be father

48
Q

how is genetic fingerprinting used to determine genetic variability? (in a population)

A

the more bands that don’t match = less related (more genetically different)

can find out how genetically varied a population is
the more the number of repeats varies at different places = more genetic variability

49
Q

uses of genetic fingerprinting (5)

A
  • determining genetic relationships
  • determining genetic variability
  • forensic science
  • medical diagnosis
  • plant and animal breeding
50
Q

what is a primer?

A

short sections of single stranded DNA
base sequence is complementary to DNA fragment so it binds
allows polymerase to bind and produce the new fragments

51
Q

why is mRNA used/ bacterial DNA better

A

mRNA and bacterial have introns removed

means bacterial DNA easier to study

fewer non-coding regions (introns)
so knowledge of genome more easily related to proteome
- less non-coding to get in the way

52
Q

promotor region

A

tells RNA polymers where to start transcription of DNA
means gene transcribed

Biology 🧬 (23 decks)

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