8. Control of Gene Expression Flashcards
1
Q
what is a gene mutation?
A
a change to the base sequence of DNA
2
Q
when are gene mutations most likely to occur
A
during DNA replication
3
Q
do gene mutations occur spontaneously?
A
yes
4
Q
what is a mutagenic agent?
A
a substance that increases the rate of mutation
5
Q
what are two different types of mutagenic agent?
A
- high energy and ionising radiation (eg gamma rays, UV and X rays)
- carcinogens eg chemicals in tobacco smoke
6
Q
what are the 6 types of mutation that can occur?
A
- substitution
- deletion
- addition
- duplication
- inversion
- translocation
7
Q
what is an addition mutation?
A
when one or more bases are added to the base sequence
eg. AGGTTAC becomes AGGTTACA
8
Q
what is the impact of an addition mutation on the base sequence?
A
all the subsequent codons are affected, resulting in a frame shift
9
Q
what is a deletion mutation?
A
when one or more bases are removed from the base sequence
eg. AGGTTA becomes AGGTT
10
Q
what is the impact of a deletion mutation on the base sequence?
A
causes a frame shift, a change to all the subsequent triplets
11
Q
what is the impact of a substitution mutation on the base sequence?
A
only one codon will change (the codon where the substitution occurred)
12
Q
what is a substitution mutation?
A
when one or more bases are swapped with eachother
eg. AGGTTAC becomes AGGTTAG
13
Q
what can mutations result in?
A
a different amino acid sequence in the encoded polypeptide
14
Q
what is an inversion mutation?
A
when a section of bases detach from the DNA sequence, and reattach at the same position but in the reverse order
eg. AGGTTAC becomes AGGATTC
15
Q
what is the impact of an inversion mutation on the base sequence?
A
new amino acids may be coded for in the affected region
16
Q
what is a translocation mutation?
A
when a section of bases detaches from the DNA sequence on one chromosome and attaches onto a different chromosome
17
Q
what are the impacts of a translocation mutation?
A
can change the resulting phenotype
18
Q
what can mutations result in?
A
a different amino acid sequence in the encoded polypeptide
19
Q
what can a different amino acid sequence in the encoded polypeptide do to the tertiary structure of a protein?
A
- the hydrogen, ionic and disulphide bonds between R groups will form in different places, causing the tertiary structure to fold differently, forming a different 3D shape and thus a non-functioning protein (or non complimentary enzyme due to change in active site shape, so no EZ complexes can form)
20
Q
why might a mutation not always affect the order of amino acids?
A
- due to the degenerate nature of the genetic code (more than one triplet of bases codes for the same amino acid), some mutations may still code for the same amino acid (eg. substitutions), so there will be no overall effect on the encoded polypeptide.
21
Q
what types of mutation will almost always cause a change to the amino acid sequence of a polypeptide? why?
A
additions, duplications and deletions
- because these mutations change the number of bases in the DNA code. this causes a frame shift in all of the triplets of bases that follow, so that each subsequent triplet is read in a different way.
22
Q
what are stem cells?
A
stem cells are unspecialised cells that continually divide/differentiate to become any type of cell (specialised)
23
Q
what is differentiation?
A
the process by which cells become specialised
24
Q
what are the four different types of stem cell?
A
totipotent
multipotent
pluripotent
unipotent
25
which type of stem cell can divide to produce any type of body cell?
totipotent stem cells
26
- for how long do totipotent stem cells occur?
- during this time, where do they occur?
- a limited time
- early mammalian embryos (hence why they are sometimes called embryonic stem cells)
27
where are pluripotent stem cells found?
in embryos
28
what do totipotent stem cells do during development?
translate only part of their dna, leading to specialisation.
29
where are multipotent and unipotent stem cells found?
in mature mammals
30
what can pluripotent stem cells do?
divide in unlimited numbers and develop into most types of body cell (cannot divide into placenta cells)
- can be used to treat human disorders
31
what can multipotent stem cells do?
divide to produce a limited number of different cell types
32
what are the issues and benefits of using pluripotent stem cells to in treatments?
- sometimes the treatment doesnt work
- the stem cells may continually divide to create tumours
- ethical debates g. whether it is right to make a therapeutic clone of yourself in an embryo to get the stem cells, and then destroy the embryo.
- stem cells can be used to grow organs that can save lives
- bone marrow cells already being used to treat leukemia
33
what can unipotent stem cells do?
can only differentiate into one type of cell, such as cardiomyocytes (heart muscle cells)
- cannot regenerate
34
what can be used to overcome the ethical issues of using embryonic stem cells? how are they produced?
- induced pluripotent stem cells (iPS)
- can be produced by taking somatic (normal) adult specialised cells and infecting them with a modified virus with genes coding for transcription factors so that the cells become pluripotent. the transcription factors will then attach to the promoter region of DNA and stimulate RNA polymerase to stimulate transcription
35
what are the sources of stem cells in mammals?
- embryos up to 16 days after fertilisation contain pluripotent stem cells
- umbilical cord contain multipotent stem cells
- the placenta has multipotent stem cells
- bone marrow contains multipotent stem cells
36
what is epigenetics?
a heritable change in gene function, without changing the DNA base sequence
37
how does increased methylation inhibit transcription?
- methyl groups bind to dna, causing dna to become more tightly wrapped around histone proteins
- this makes the dna less accessible to transcription factors, so transcription factors cant bind to dna, and transcription does not occur.
38
how does decreased acetylation of histones inhibit transcription?
- decreased acetylation makes the histones more positive (as they are losing negative charge), so the dna and histones no longer repel, and become more closely associated with eachother
- this makes it harder for transcription factors to bind, so transcription does not take place
39
how could the hypermethylation of tumour supressor genes cause cancer?
methyl groups added to a tumour supressor gene, so tumour suppressor gene is not transcribed, leading to uncontrollable cell division
40
what is a protooncogene?
a protein that stimulates cells to divide
41
what is an oncogene?
a mutated proto-oncogene, resulting in the gene constantly being switched on, leading to uncontrollable cell division
42
how could the hypomethalation of proto-oncogenes cause cancer?
- too little methyl groups being attached
- so the proto-oncogene is easier to transcribe, and is always switched on
- begins to behave like an oncogene
- leading to uncontrollable cell division
43
what is a transcription factor?
a protein that controls the rate of transcription of target genes
44
how do transcription factors work?
they bind to specific dna base sequences and either help rna polymerase to bind, or inhibit it
- if they help it, then rna polymerase can copy the dna base sequence onto an mrna strand, which can then move into the cytoplasm
- if it doesnt help it, the gene will be turned off
45
what is a steroid hormone? give an example
- a molecule that binds to transcription factors and activates them
- oestrogen
46
explain how oestrogen can initiate transcription
- small and hydrophobic so can diffuse through the phospholipid bilayer into cells
- it binds to the complementary receptor of a transcriptional factor
- when it binds, it changes the shape of the dna binding site slightly, thus activating the transcription factor
- this change in shape of the dna binding site makes it complementary to the dna, so it can bind to dna, and only then can RNA polymerase attach and form mRNA
47
how can the translation of mrna be inhibited?
what types of organisms experience this?
- rna interference
- eukaryotes, and some prokaryotes
48
describe the process of rna interference
- small interfering RNA (siRNA) is double stranded RNA
- this double stranded siRNA associates with enzymes to become single stranded and cut into small sections
- siRNA binds to another enzyme in the cytoplasm and this siRNA-enzyme complex binds to an mRNA molecule by complementary base pairing
- the enzyme associated with siRNA then cuts the mrna into fragments, so it can no longer be translated
49
describe how miRNA can inhibit the translation of mRNA
- in mammals, miRNA isnt fully complementary to target mRNA, making it less specific than siRNA so it may target more than one mRNA molecule
- miRNA associates with an enzyme and binds to target mRNA i the cytoplasm
- miRNA blocks the translation of the target mRNA
- mRNA then moved to a processing body where it is stored or degraded
50
what is the development of cancer the result of?
mutations in the genes that regulate mitosis
51
what is a tumour?
a mass of unspeciallised cells formed from uncontrolled cell division
52
what are the two types of tumour?
benign
malignant
53
what are benign tumours?
non-cancerous tumours that grow at a slower rate and do not invade neighbouring tissues
54
why do benign tumours not invade neighbouring tissues?
- produce adhesion molecules which stick them together and to a particular tissue
- often surrounded by a capsule
- remain compact and do not metastasise
55
how can a benign tumour cause harm?
- cause blockages/obstructions
- may damage organ concerned
- may put pressure on other organs
56
what are malignant tumours?
cancerous tumours that grow rapidly and invade neighbouring tissues
- break off and form new tumours, spreading to other parts of the body
- can develop their own blood supply
57
how is oestrogen linked to the development of breast cancer?
- increased exposure to oestrogen due to starting period early or starting menopause late
- after menopause, oestrogen no longer produced in ovaries, but instead in the fat cells in breast tissues
- oestrogen can stimulate breast cells to divide and replicate which increases the chances of mutations occurring
58
what is the genome?
the entire set of genetic material of an organism
59
what is meant by sequencing a genome?
working out the dna base sequence for all the dna in a cell
60
what does determining the genome of simpler organisms allow?
the identification of the proteome
61
what is the proteome?
the full range of proteins a cell is able to produce / the number of different proteins in the genome
62
why is it easier to determine the proteome of prokaryotic organisms?
because they do not contain introns (non coding regions of dna)
63
what is the identification of the proteome useful for?
help identify the potential antigens to use in a vaccine
64
why is the genome of complex organisms harder to sequence/translate into the proteome?
because they contain large sections of non coding dna (introns) and regulatory genes
65
state two features of current sequencing methods:
- continuously updated
- have become automated
66
what does recombinant dna technology involve?
why can the transferred dna be translated within the cells of the recipient organism?
the transfer of fragments of DNA from one organism or species to another
because the genetic code is universal, as well as transcription and translation mechanisms
67
what are the organisms that receive the dna fragments from another organism during recombinant dna technology called?
transgenic organisms
68
what are the three methods of obtaining fragments of dna for recombinant dna technology?
- conversion of mRNA to complimentary DNA using reverse transcriptase
- using restriction enzymes to cut a fragment containing the desired gene from DNA
- creating the gene in a gene machine
69
describe the process of using restriction enzymes to create dna fragments for recombinant dna technology
- mrna is isolated from the cell
- reverse transcriptase joins DNA nucleotides lined up opposite the mrna strand TO the mrna strand using complimentary base pairing
- single stranded, complimentary dna (cDNA) is made
- to make fragment double stranded, DNA polymerase is used
70
advantages of using reverse transcriptase to make dna fragments?
- the cdna produced is intron free as its based on mrna template
- mrna present in large amounts in cell making the protein
71
what are restriction endonucleases?
enzymes that recognise specific palindromic sequences and cut dna at these places
72
what is the area a restriction enzyme is able to cut called?
the recognition site - this is the area it is complimentary to
73
why is it preffered when the restriction endonuclease enzymes cut staggered palindromic ends?
because they contain exposed dna bases called 'sticky ends' which can be aligned with the exposed bases of the receiving organism's dna, making the dna easier to anneal to any other fragment cut using the same restriction enzyme (as the sticky ends produced would be complimentary)
74
advantages and disadvantages of using restriction endonucleases to create dna fragments
adv = sticky ends easier to insert to make recombinant dna
dis= still contains introns
75
describe how dna fragments can be produced using a gene machine
- the sequence that is required is designed (if it doesnt already exist)
- the first nucleotide in the sequence is fixed to a support eg a bead
- nucleotides are added one by one in the correct order, including protecting groups to ensure the nucleotides are joined at the right points to prevent unwanted branching
- short sections of dna called oligonucleotides are produced. then they are broken off from the support and the protecting groups are removed
- oligonucleotides can then be joined together to make longer dna fragments
76
advantages of using gene machine to create dna fragments
- very quick and accurate
- makes intron free dna (so can be transcribed in prokaryotic cells)
- can design exact dna fragment wanted
77
disadvantage of using gene machine to create dna fragments
need to know the sequence of amino acids or bases
78
what does it mean to amplify dna fragments??
to clone them to create large numbers
79
what are the two ways dna fragments can be amplified?
in vivo = using vectors
in vitro = polymerase chain reaction (PCR)
80
what is needed for PCR to take place?
- primers
- dna nucleotides
- dna polymerase
- dna fragment to be amplified
81
what are primers?
short lengths of single stranded dna that are complementary to the bases at the start and end of the fragment you want
82
why is dna polymerase taken from extremophiles used?
- has evolved to have a higher optimum than normal dna polymerase
- will not denature with high temperatures used in PCR
83
describe the process of PCR
- requires dna fragment, dna polymerase, dna nucleotides and primers
- heat to 95C to break hydrogen bonds and separate DNA strands which become templates
- decrease temperature to 55C to allow primers to bind to DNA strands
- increase temperature to 70C, which is the optimum for DNA polymerase, so DNA polymerase can join nucleotides by complimentary base pairing after attaching to primers
- repeat cycle to create many copies
84
how many new copies of DNA are made in one cycle of PCR?
2
85
each PCR cycle ___ the amount of strands
doubles
- first cycle produces 2x2= 4 dna strands
- second cycle produces 4x2=8 strands etc
86
what are the advantages of using PCR ?
- its automated making it more efficient
- its rapid
- it doesnt require living cells, so less complex techniques needed
87
what is a vector?
what are the most common vectors?
something that carries the isolated dna fragment into the host cell
plasmids
88
what is a promoter reigon?
a dna sequence added at the start of a fragment that binds to rna polymerase to enable transcription to occur
89
what is a terminator reigon?
dna sequence added at the end of a fragment that causes rna polymerase to detach and stop transcription
90
describe the process of in vivo amplification
- add terminator and promoter regions
- restriction endonuclease cut dna fragment to produce sticky ends
- the same restriction endonuclease enzyme used to cut open the vector (plasmid), to produce complementary sticky ends
- dna ligase used to anneal the dna fragment and the plasmid (now called recombinant dna)
- add marker gene
- the recombinant dna is then inserted into the host cell where the gene will be expressed (the protein it codes for will be made)
91
why might the recombinant dna not be successfully inserted into the host cell?
- recombinant plasmid may not enter the host cell
- plasmid re-joins before dna fragment is inserted
- dna fragment sticks to itself, rather than inserting into the plasmid
92
how can the host cells that succesfully take up the recombinant dna be identified?
with marker genes
93
what are three common types of marker gene?
- antibiotic resistance genes
- genes coding for flourescent proteins
- genes coding for enzymes
94
what are dna probes and dna hybridisation used for?
to locate specific alleles of target genes
95
what is a dna probe?
a short single strand of DNA that has bases complimentary to the alleles of genes
96
how do dna probes work?
- patients dna is treated to make it single stranded
- patients dna is mixed with dna probes
- if the patient has the allele, dna probe will hybridise (bind) to the target allele and a label will indicate its presence
97
describe the process of dna hybridisation
- patients dna is heated to make it single stranded (hydrogen bonds between bases break)
- patients single stranded dna is mixed with dna probe, and mixture is cooled so that complimentary base pairs can align and anneal
98
what must be known before a dna probe can locate a specific allele?
the dna base sequence
99
why would the dna mixture be washed after the hybridisation step?
to remove any unbound dna probes
100
what are the uses of dna probes?
- screen patients for heritable conditions
- screen patients for drug responses
- screen patients for health risks
101
what is the purpose of a dna microarray?
multiple different dna probes screening for multiple different diseases / genetic disorders
102
what is genetic counselling?
advising patients and their relatives about the likelihood of them carrying any alleles linked to diseases or genetic disorders
103
uses of genetic counselling
- screening for conditions that can be treated early on eg breast cancer gene could lead to masectomy
- deciding not to screen for inevitable conditions that may affect how the patient lives their whole life eg. huntingtons
104
what are personalised medicines?
medicine tailored to an individuals dna, to predict how people will respond to different drugs
105
- what does VNTR stand for?
- where can they be found?
- what are VNTR'S
- variable number tandem repeats
- in an organisms genome (within introns)
- non coding sequences of dna
106
what is the probability of 2 individuals having the same VNTRS's
very low
107
what is the purpose of genetic fingerprinting?
analysing dna fragments that have been cloned by PCR, determining genetic relationships, and the genetic variability within a population
108
describe the process of genetic fingerprinting
- sample of dna is obtained eg. from the blood, saliva etc
- pcr is used to make many copies of the dna sample
- restriction endonucleases complimentary to sequences closer to the VNTRs added to cut the dna into smaller fragments
- flourescent dna probe is added to dna samples so they will be visible under uv
- dna samples are placed into gel electrophoresis, which separates samples according to length/mass
- make dna single stranded
- transfer to nylon membrane
- dna fragments viewed as bands under uv light
- position of dna bands are compared
109
what are the uses of genetic fingerprinting?
- forensic science = place suspects at crime scenes
- medical diagnosis
- animal and plant breeding (ensure they are not closely related before breeding)
110
describe the process of gel electrophoresis
- dna mixture placed into a well in a slab of gel covered in buffer solution that conducts electricity
- current passed through the gel, and dna fragments are negatively charged (due to phosphate group) so move towards the positive electrode at the farther end of the gel
- small dna fragments move faster and travel further through the gel, so dna fragments separate out according to size/length
