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Biology OCR A Alevel > Manipulating Genomes > Flashcards

Manipulating Genomes Flashcards

1
Q

What are the 5 components needed in DNA sequencing?

A

DNA for sequencing

DNA primer

DNA polymerase

Normal bases

Terminator bases (with fluorescent tag)

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2
Q

What does DNA sequencing do?

A

Determined the precise order of nucleotides within a DNA molecule

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3
Q

What are the stages of DNA sequencing?

A

Mix the DNA to be sequenced with the dna primer, DNA polymerase and excess normal and terminator bases

Place in a thermal cycle

95 degrees - break hydrogen bonds and separate strands

50 degrees for primers to anneal

60 degrees for DNA polymerase to work best and add nucleotides to complementary bases

Each time terminator base is added, the chain stops producing many different lengths chains

Separated by electrophoresis or capillary sequencing when fluorescent markers on terminators are used to identify the final base on each fragment

Order of bases produced shows sequence of the complementary strand to the dna sample to work out the original from

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4
Q

What are bioinformatics?

A

Software and computing tools to organise and analyse raw biological data

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5
Q

What is computational biology?

A

Use of data to build theoretical models of biological systems and predict what could happen under certain conditions

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6
Q

What are 3 things bioinformatics and computational biology contribute into the research of?

A

Genotype-phenotype relationships

Epidemiology

Evolutionary relationships

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7
Q

What are 3 things gene sequencing has allowed to happen?

A

Comparison between individuals and between species

Sequences of amino acids in polypeptides to be predicted

Development of synthetic biology

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8
Q

What is the synthetic biology?

A

The design and construction of novel artificial biological pathways, organisms or devices

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9
Q

What is DNA profiling?

A

Produce an image of the patterns in the non coding satellite regions of introns of DNA

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10
Q

How is DNA profiling done?

A

Extract DNA and perform PCR

Digest sample with restriction endonuclease enzymes

Separate strands by electrophoresis

Hybridisation - adding radioactive or fluorescent probes (short complementary sequences to known sequences)

X-ray or apply UV light depending on probe used to see evidence

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11
Q

What is DNA profiling used for?

A

Forensics
Comparing dna sample found at crime scene to suspects

Analysis of disease risk
Some micro satellite regions are associated with increased risk of disease, specific gene markers can be identified and observed in DNA profiles to assess the risk

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12
Q

What are the 3 stages of PCR

A

Separate the strands of DNA by heating to 95 degrees which breaks the hydrogen bonds between bases

Annealing primers, cool to 55 degrees so primers anneal to ends of the strands

Heat to 72 degrees optimum temperature for taq polymerase and bases are added to the primer making complementary strand

Repeat

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13
Q

Why is taq polymerase used?

A

Taq polymerase is a bacterial DNA polymerase and it has much higher optimum temperature that human DNA Polymerase so does not denature when heated during PCR

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14
Q

What are the uses of PCR?

A

DNA Profiling

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15
Q

What are the three stages of electrophoresis?

A

Load DNA fragments into the wells of the agarose gel strips which contain a buffer solution to maintain constant pH

Electric current is passed through electrophoresis plate making fragments at cathode end move towards the anode due to the negative phosphate group

The smaller fragments move further and faster as mesh structure of gel resists the movement of the larger fragments when smallest has reached the end the current is switched off

Southern blotting - alkaline buffet solution is added to denature the fragments and separate the strands. Nylon membrane is placed on top the many dry absorbent paper sheets this draws up alkaline solution by capillary action but dna fragments can’t get through so transferred to the membrane the fixed by UV light or 80 degrees

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16
Q

What do you change in electrophoresis when applying it to a protein?

A

heat it to denture the protein to expose the hydrophobic region/ expose charges

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17
Q

name 4 purposes of a modern botanic garden

A

conservation of rare endangered species

acts as a gene bank

for teaching/education

as visitor attraction- aesthetic reasons/leisure

18
Q

what is the role of PCR in sequencing genomes?

A

to amplify and make many copies of DNA

which will be of different lengths

19
Q

what is the role of electrophoresis in sequencing genomes?

A

to put DNA pieces in size order ;

to read, base sequence / order of bases ;

20
Q

what is the role of restriction enzymes in sequencing genomes?

A 
to cut (genome DNA) into, small(er) / 750 bp, fragments ;
to cut, vectors / BACs / plasmids, (for gene library) ;
21
Q

why does a genome have to be fragmented before sequecning?

A

genome, too big / very large ;
accuracy better / fewer errors (with small fragments) ;
divide job over, time / different labs ;

22
Q

what can be used as a vector for bacteria?

A

plasmid

BAC

bacteriophage

23
Q

what can be used as a vector for plants?

A

plasmid

virus

argobacterium tumefaciens

24
Q

what can be used as a vector animals?

A

virus

plasmid

BAC

25
Q

state the 3 main stages of genetic engineering

A

isolation of the desired gene

inserting gene into a vector

inserting vector into the organism

26
Q

describe how the gene is isolated in genetic engineering

A

by restriction endonuclease enzymes
at specific base sequences - palindromic sequences

recognition site is complementary to the enzyme

leaves sticky ends which are unpaired bases at each end of the fragment or leaves blunt ends

27
Q

describe how vectors are used in genetic engineering

A

the same restriction endonucleases used to cut the DNA fragment are used to cut the vector - for example a plasmid

this means the sticky ends on the vector are complementary to the DNA fragment containing the gene

the vector DNA and DNA fragment are mixed together with DNA ligase which joins the sugar phosphate backbone.

forms recombinant DNA

28
Q

what can be done to encourage the cell to take up the vector e.g. the plasmid?

A

electroporation

bacteria cells are mixed with the plasmid vector and placed in electroporator

an electric feild is applied

this increases permiability of bacterial cell membrane which allows them to take in the plasmid

29
Q

give an example of genetic manipulation in microorganisms

A

genetically modified pathogens for research

30
Q

give an example of genetic manipulation in plants

A

genetically modified soya for insect resistance

31
Q

give an example of genetic manipulation in animals

A

pharming- genetically modified animals to produce pharmocueticals

32
Q

what is gene therapy?

A

altering alleles inside cells to cure genetic disorders using vectors such as viruses, plasmids and liposomes

33
Q

what are the 2 types of gene therapy?

A

somatic and germ line

34
Q

what is somatic gene therapy?

A

altering body cells , particularly those that will be most affected

35
Q

what is germ line gene therapy?

A

altering the alleles in germ cells - sex cells or early embryo so every subsiquent cell of organism is affected

36
Q

what are positive and negatives of somatic gene therapy?

A

pros
cure / reduce symptoms / better quality of life / less medication;
cystic fibrosis / SCID / Parkinson’s / thalassaemia / LCA ;
extend lifespan / saves lives ;

cons
virus vector may cause (viral) disease ;
procedure may be, invasive / dangerous / painful / stressful ;
temporary / needs to be repeated / limited success ;
immune system / rejection, problems ;
animal testing concerns ;

37
Q

what are the negative ethical issues of gene therapy?

A

they could be used in non medical ways- for cosmetic reasons

could do more harm than good - risk of overexpression of the gene

it is expensive so questionable wether this is the best way health services could spend their resources

38
Q

what are the 5 main stages of genome sequencing?

A

extract samples of DNA from cells

cut DNA into sections of varying lengths

amplify the DNA

sequence short sections of DNA

place sections in order by matching overlapping regions

39
Q

what is electrofusion?

A

another way of producing genetically modified cells

tiny electric currents are applied to the membranes of 2 different cells, this fuses the cell and nuclear membrane of the 2 cells to form a hybrid or polyploid cell containing DNA from both-
used successfully to produce GM plants

40
Q

how can bioinformatics be useful in determining wether a newly sequenced allele causes genetic disease

A

base sequence of normal allele and (known) alternatives
held (in database) 
computational analysis allows rapid comparison of
sequences with newly sequenced allele 
amino acid sequence / protein structures, also held
(in database) 
idea of
computer modelling of new protein structure from base
sequence 

41
Q

why are only non coding sections of DNA selected for profiling a human?

A

in most people, the genome is very similar / most genes the
same 
using coding sequences would not provide unique profiles 
(parts of) non-coding DNA contains variable numbers of ,
short tandem repeats / repeating sequences 

42
Q

why are proteins heated before being placed in electrophoresis gel?

A

to denature the protein AND exposes charges or hydrophobic region

Biology OCR A Alevel (26 decks)

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