SECTION 8 - GENE EXPRESSION, RECOMBINANT DNA TECH.

Question 1

mutation

Answer 1

a change in the dna base sequence

Question 2

mutation can occur in 2 places. where?

Answer 2

1) in somatic cells (i.e body cells), which may result in cancer
2) in gametes, which means the mutation will be passed on to the offspring if the cell with the mutation takes part in fertilisation.

Question 3

SUBSTITUTION MUTATION

Answer 3

1 BASE IS SUBSTITUTED FOR ANOTHER

Question 4

DELETION MUTATION

Answer 4

1 OR MORE BASES ARE LOST

Question 5

ADDITION MUTATION

Answer 5

1 OR MORE BASES ARE ADDED

Question 6

DUPLICATION MUTATION

Answer 6

A SEQUENCE OF BASES OR A WHOLE GENE IS INSERTED TWICE OR MULTIPLE TIMES. THIS IS IMPORTANT IN EXPANDING THE GENOME AS ONE OF THE GENES CAN MUTATE AND CODE FOR A NEW POLYPEPTIDE, WHILE THE ORIGINAL CONTINUES TO WORK AS NORMAL.

Question 7

inversion mutation

Answer 7

a base sequence is removed, rotated by 180 degrees, and inserted back again. very harmful as almost every codon differenent and polypeptide produced is dysfunctional.

Question 8

TRANSLOCATION MUTATION

Answer 8

a sequence of bases is taken out and inserted at a different position in the genome. may be within the same chromosome or between different chromosomes.

Question 9

why might a mutation not have an effect upon the phenotype?

Answer 9

some take place within introns
some take place in non-coding regions between genes
cuz the genetic code is degenerate, the sequence of encoded amino acids may still be the same as the new codon may code for the same amino acid. may change the amino acid but no effect teriary structure as same bonds formed. ( so a sub mutation may not be harmful)

Question 10

a sub mutation may not be harmful as it doesnt cause a frameshift. the rest are very harmful. mutations like addition, deletion and duplication cause a frameshift. whats that

Answer 10

when all bases shift in one direction because the number of bases is changed, hence every codon is read differetnly

Question 11

what are mutagenic agents, and state the effects of each of the types.

Answer 11

agents that increase the rate of mutation.
Base Analogs - can substitute for a base during DNA replication, which changes the base sequence in the new DNA.
some chemicals - may delete/alter bases.
Radiation - may alter the structure of DNA, leading to problemds during replication.

Question 12

give the 2 key properties of stem cells

Answer 12

1) the ability to divide and replicate
2) potency (the ability to become specialised)

Question 13

TOTIPOTENT STEM CELLS

Answer 13

THESE ARE STEM CELLS THAT CAN DIVIDE AND DIFFERENTIATE INTO ANY TYPE OF BODY CELL
THEY ONLY OCCUR FOR A VERY LIMITED TIME IN EARLY MAMMALIAN EMBRYOS

Question 14

DURING DEVELOPMENT WHAT DO TOTIPOTENT CELLS DO? ( TO RESULT IN CELL SPECIALISATION)

Answer 14

THEY TRANSLATE ONLY PART OF THE DNA, RESULTING IN CELL SPECIALISATION

Question 15

PLURIPOTENT

Answer 15

THESE STEM CELLS ARE DESCENDANDS OF TOTIPOTENT CELLS AND CAN DIFFERENTIATE INTO ANY TYPE OF BODY CELL EXCEPT THOSE THAT MAKE UP THE PLACENTA.

Question 16

MULTIPOTENT

Answer 16

MULTIPOTENT CELLS ARE ONES THAT CAN DIFFERENTIATE INTO A LIMITED NUMBER OF TYPES OF BODY CELLS

Question 17

UNIPOTENT

Answer 17

UNIPOTENT CELLS ARE CELLS THAT CAN DIFFERENTIATE INTO ONLY 1 TYPE OF CELL

Question 18

WHICH 2 TYPES OF STEM CELLS ARE FOUND IN MATURE MAMMALS

Answer 18

UNI AND MULTIPOTENT

Question 19

TOTI POTENT VS PLURIPOTENT DIAGRAM

Answer 19

https://merchanttaylorsschools-my.sharepoint.com/:w:/g/personal/tofoma_merchanttaylors_com/Eawz4omNH7NAgYW6F-uVzuEBHGEeiQ36q8dV4BDgdp01zg?e=wZdu8f

Question 20

USE OF PLURIPOTENT CELL

Answer 20

they can divide in unlimited numbers and can be used to treat human disorders (eg grow replacement tissues in a lab, replace damage tissues)

Question 21

USE OF unipotent cardiomyocytes (heart muscle cells)

Answer 21

COULD DEVELOP INTO NEW HEART TISSUES

Question 22

WHAT are induced pluripotent stem cells (iPS cells), why are they made and whats the advantage of using them

Answer 22

induced pluripotent stem cells (iPS cells) are a type of pluripotent cell that are made by reprogramming adult somatic (body) cells. They are made to express a series of transcription factors that are normally associated with pluripotent stem cells, meaning that genes are expressed that are associtated with pluripotency.
the adv’s of ips cells: (no harvesting of embryonic stem cells), so no destruction of embryo, meaning ethical issues are avoided.
no risk of rejection by the patient as the ips cells are made using the patients own cells

Question 23

give 2 issues surrounding the use of stem cells

Answer 23

1) the ethical issue of the fact that an embryo has the ability to form a human
2) stem cells can multiply out of control in the body forming tumours

Question 24

TRANSCRIPTION FACTOR

Answer 24

A transcription factor is a protein that helps control the process of transcription, which is the first step in gene expression. TRANSCRiption factors move from the cytoplasm into the nucleus. here they bind to specific promoter regions near the start of their target genes

Question 25

what are the 2 types of transcription factors and what do they do

Answer 25

Activators, which increase the rate of transcription by making it easier for RNA polymerase to bind to the start of the target gene. Repressors, which decrease the rate of transcription by making it harder for RNA polymerase bind to the start of the target gene. note start of gene = promoter region https://merchanttaylorsschools-my.sharepoint.com/:w:/g/personal/tofoma_merchanttaylors_com/ESBehuwbziVJgzfEVl5fHvMBriRt646OEPcAFfJT-SHjZQ?e=0Er41X

Question 26

THE ROLE OF OESTROGEN IN INITIATING TRANSCRIPTION. (include why they are able to diffuse through the phospholipid bilayer, what it binds to, etc)

Answer 26

Oestrogen is a steroid hormone. steroids are lipids (lipids are non polar hence can diffuse via the hydrophobic core of the phospholipid bilayer) , so can diffuse through the phospholipid bilayer of cell surface membranes, entering the cytoplasm of the cell. in the cytoplasm, oestrogen binds to an oestrogen receptor (ERa) to form an oestrogen-oestrogen receptor complex. This leaves the cytoplasm and passes into the nucleus were it acts as a transcription factor which binds to the promoter region of a specific gene. NOte that the oestrogen-ERa compex acts as a transcription factor for many genes and can both activate and repress different genes.

Question 27

EPIGENETICS

Answer 27

EPIGENETICS INVOLVES HERITABLE CHANGES IN GENE FUNCTION WITHOUT CHANGES TO THE BASE SEQUENCE OF DNA. THESE CHANGES INSTEAD ARE CAUSED BY CHANGES IN THE ENV, THAT INHIBIT TRANSCRIPTION BY: 1) INCREASED METHYLATION OF DNA. where a methyl group becomes attached to the DNA coding for a gene. the methyl group always attached at a CpG site (where a cytosine and guanine base are next to each other in the sequence). the binding of the methyl group changes the DNA's structure, making it harder for transcription factors/RNA polymerase to bind to promoter regions and hence transcribe DNA. 2) Decreased acetylation of assosciated histones: DNA is wound around histones to form chromatin. if acetyl groups bind to chromatin, this reduces the attraction between DNA and histones, leaving chromatin to be 'relaxed' (open), making it easier for transcriptional machinery to bind to DNA, which increases rate of transcription. if acetyl groups are removed from histones, the opposite happens, as attraction between dna and histones is increased and hence chromatin is said to be closed, making it harder for transcription machinery to bind to dna, decreasing the rate of transcription REMEMBER TO ALWAYS SAY ACETYLATION OF ASSOSCITED HISTONES NOT ACETYLATION ON ITS OWN. decreased acetylation of associated histones - https://merchanttaylorsschools-my.sharepoint.com/:w:/g/personal/tofoma_merchanttaylors_com/Ea7f_vHft5lDveszeeu9OqQBZqHgpwveBzg8p9yOZ68ZZQ?e=Lq3yS4

Question 28

note that there are lots of cpg sites found in promoter regions. what does this mean for the gene

Answer 28

it may be silenced

Question 29

when can epigenetic changes be inherited by offspring?

Answer 29

if they occur in gametes.

Question 30

in eukaryotes and some prokaryotes how can the translation of mrna produced from target genes be inhibited(prevented) by RNAi (RNA interference)?

Answer 30

small interfering RNA (siRNA) is a short double stranded RNA molecule that assosciates with proteins whereby: the double strand is separated into 2 strands one of these siRNA strands combines/binds with a protein the RNA-protein complex binds to an mrna strand that is complementary to it the protein cuts the mrna into fragments, and finally a processing body degrades (breakdsdown) the fragments, as a result the mrna is not translated. https://merchanttaylorsschools-my.sharepoint.com/:w:/g/personal/tofoma_merchanttaylors_com/EYYAuHNHaU1Pkea0ZDspA7QBUN6wQcfWzONZrgX5ou9Rvw?e=pWnwno

Question 31

GIVE THE 2 TYPES OF TUMOUR AND DIFFERENTIATE BETWEEN THEM.

Answer 31

https://merchanttaylorsschools-my.sharepoint.com/:w:/g/personal/tofoma_merchanttaylors_com/EXqR_wmsWYVNoV6Ip3I1VJgBm7pUVTmiLvI3h_AXeBMvWw?e=0vgTjK

Question 32

give the 2 types of genes that cntrol cell division

Answer 32

tumour suppresor genes proto-oncogenes/oncogenes

Question 33

What do tumour suppressor genes do, and how can their malfunction lead to cancer?

Answer 33

They code for proteins that stop cell division or cause apoptosis (i.e cell death) If a mutation occurs in a tumour suppresor gene, cells divide uncontrollably. Hypermethylation can silence these genes, so their proteins which they code for arent made/synthesised, and hence more cell division takes place.

Question 34

What do proto-oncogenes do, and how can they lead to cancer?

Answer 34

They code for proteins that stimulate cell division. If a mutation occurs in a proto-oncogene, they can become overactive oncogenes, causing uncontrolled cell division. Hypomethylation can lead to too much expression of these genes leading to increased production of proteins, encouraging excessive division.

Question 35

lifestyle can have a large impact on the likelihood of cancer developing, as it can influence on epigenetic processes like....

Answer 35

methylation

Question 36

NOTE THAT EVEN IF MUTATIONS OCCUR, A TUMOUR MAY BE PREVENTED FROM FORMING, IF WHAT?

Answer 36

1) repair mechanisms for dna work 2) cells with the mutation may be detected by the immune system and destroyed

Question 37

How is oestrogen levels (concs) linked to an increased risk of cancer?

Answer 37

Increased oestrogen levels (concs) may cause increased activation of proto-oncogenes. This can lead to uncontrolled cell division and tumour growth.

Question 38

Why can oestrogen increase the risk of cancer after menopause?

Answer 38

Fat cells produce more oestrogen after menopause → [oestrogen] rises. More dividing cells = more oestrogen produced = tumour grows faster. White blood cells attracted to tumour cells also increase oestrogen. Oestrogen may even cause mutations in the DNA of breast cells.

Question 39

GENOME

Answer 39

THE COMPLETE SET OF GENETIC MATERIAL OF AN ORGANISM (so includes mitochondrial and chloroplast DNA). it is All dna in an organism

Question 40

sequencing (genome) projects have read the genomes of a wide range of organisms. hence they can be used to determine what between organisms?

Answer 40

since they have read the genomes of a wide range of organisms, they can use that to determine evolutionary relationships between organisms.

Question 41

simple organisms like bacteria and fruit flies etc, do not have ....... meaning all their dna is coding

Answer 41

they dont have introns

Question 42

why is the dna in simple organisms more accessible, and hence since its more accessbile we can easily determine the....., and why? and whats the advantage of determining this (thing) ? (USE OF GENOME PROJECTS)

Answer 42

cuz it is shorter and has no histones in simple organisms. this means that we can easily determine the proteome (full set of proteins/sequence of proteins that derive from the genetic code) from the genome, (because its the dna base sequence which determines the sequence of amino acids in a specific polypeptide.) DETERMINING THE PROTEOME FROM THE GENOME MEANS (adv): IT ALLOWS FOR THE IDENTIFICATION OF POTENTIAL ANTIGENS ON THE SURFACE OF BACTERIA (OR VIRUSES - 'GENOME' CAN ALSO BE USED TO REFER TO VIRAL RNA) TO HELP DEVELOP VACCINES. (EG, MRNA VACCINES)

Question 43

UNLIKE IN SIMPLER ORGANISMS, WHY IS GENOME SEQUENCING MORE DIFFICULT IN COMPLEX ORGANISMS

Answer 43

COMPLEX ORGANISMS CONTAIN: PRESENECE OF INTRONS AND NON CODING MULTIPLE REPEATS PRESENCE OF REGULATORY GENES (which control which genes are translated or not), meaning we have to identify the regions which are coding and non-coding first. knowledge of the genome can therefore not be easily transported to the proteome. note that sequencing methods are continously updated and have become automated and cheaper

Question 44

What is recombinant DNA technology and why can it work across different species?

Answer 44

It involves transferring fragments of DNA from one organism (or species) to another. It works across diff species because the genetic code is universal and hence the mechanisms for transcription and translation are the same. This allows the transferred DNA to be translated inside the recipient (transgenic) organism.

Question 45

how can u produce dna fragments via using reverse transcriptase?

Answer 45

using reverse transcriptase: mrna for a specific polypeptide is isolated from the cell the mrna is then mixed with free dna nucleotides and reverse transcriptase the reverse transcriptase alongisde a dna molecule then uses the mrna as a template to produce cDNA (complementary DNA), which is a double-stranded copy of the require gene https://merchanttaylorsschools-my.sharepoint.com/:w:/g/personal/tofoma_merchanttaylors_com/EcxNlLsf6Y1Dqx73FV9oNLwBoXHsh3g_6SDAnZo8SFnwLw?e=HeJqLk

Question 46

in the production of dna fragments via the use of reverse transcriptase, why is using mrna useful?

Answer 46

as: most cells only have 2 copies of each gene, whereas they have lots of mrna of that gene that arereadily available and hence easier to obtain. mature mrna has no introns

Question 47

What is the role of restriction endonucleases and vectors in recombinant DNA technology, and how are sticky ends involved? ( how can u produce dna fragments via restriction endonucleases enzymes)

Answer 47

Restriction endonucleases have an active site complementary to a specific recognition site on DNA. They bind to the recognition site and cut the DNA, often creating "sticky ends" (which are staggered cuts that reveal unpaired bases at either end of the DNA fragment or plasmid). A vector (such as a plasmid or bacteriophage) is used to transfer DNA into a cell. If both the vector and the DNA fragment are cut with the same restriction endonuclease, their sticky ends will be complementary. DNA ligase joins the sticky ends of the vector and fragment, forming phosphodiester bonds and creating recombinant DNA. https://merchanttaylorsschools-my.sharepoint.com/:w:/g/personal/tofoma_merchanttaylors_com/ES_o0lXUCzlNoAMzVxOquVgBx9BWFYHafbQmF3H574ih3A?e=t4JTXL

Question 48

why is using restriction enzymes less useful

Answer 48

cuz they make cut dna with introns

Question 49

how can u produce dna fragments via using a gene machine

Answer 49

determine the amino acid sequence of specific polypeptide this tells us the mrna base sequence from which we can determine the dna base sequence which acted as a template for it a machine then assembles and joins together nucleotides in the desired sequence once DNa fragments are isolated, amplify it so that you have sufficient copies to work with. amplification of these dna fragments are done in 2 types of clonings: invivo amplification and invitro amplification.

Question 50

give the whole procedure of invivo cloning

Answer 50

Transformation takes place, which involves insertion of the recombinant DNA into a host cell if a plasmid is the vector: host cells can be stimulated to take in the plasmid if bacteriophage is the vector: bacteriophage injects its dna into the bacterium Next identification takes place: only very few host cells will take up the recombinant DNA to identify which have taken up the DNA, we use marker genes which are inserted into the vector together with the required gene. marker genes may code for: code for antibiotic resistance. the bacteria can be grown on an agar plate containing the antibiotic, and so only transformed cells (which have taken in the recombinant DNA) will survive and produce colonies. code for fluorescence (and the cells which have taken in the recombinant DNA can be identified using UV light) Finally the transformed bacteria are allowed to reproduce. this is known as invivo cloning.

Question 51

in invivo cloning why must you ensure promoter and terminator regions are present in the recombinant DNA,

Answer 51

cuz promoter and terminator regions determine where dna polymerase starts and stops replication. these regions may already be present in the vector dna or they may have to be added to the dna.

Question 52

why would we use PCR

Answer 52

to test for the presence of a specifc DNA base sequence (eg, to test for viral infections) genetic fingerprinting Genetic modification to transform organisms MOST IMPORTANTNLY AND GENERALLY TO AMPLIFY DNA FRAGMENTS

Question 53

how does PCR work? (INVITRO METHOD)

Answer 53

1) A reaction mixture is set up containing: DNA fragment to be copied primers dna polymerase free dna nucleotides note that a primer is a short single stranded dna base sequence thats complementary to the bases at the start of the desired DNA fragment. it allows dna polyermase to bind to the start of the dna fragment. dna polymerase: for PCR to work, high temps are required hence taq polymerase (a type of dna polymerase obtained from thermophilic bacteria, and hence is heat resistant) is use. 2) step 2: heat reaction mixture to 95 degrees C, causing the DNA to denature, meaning H bonds between strands break. 3) step 3: cool the mixture to 50-60 degrees, allowing primers to anneal (bind) to complementary bases at ends of fragments. 4) step 4: heat to 72 degrees so that dna polymerase can carry out extensions of the strands, meaning new complementary strands are formed. repeat this process many times

Question 54

FRAGMENTS OF DNA CAN BE AMPLIFIED VIA 2 TECHNIQUES GIVE THEM

Answer 54

INVITRO TECHNIQUES AND INVIVO TECHNIQUES

Question 55

GIVE THE 3 WAYS TO PRODUCE FRAGMENTS OF DNA?

Answer 55

conversion of mrna into complementary DNA (cDNA) using reverse transcriptase using restriction enzymes to cut a fragment containing the desired gene from DNA creating the gene in a gene machine.

Question 56

DNA PROBE and WHAT IT DOES

Answer 56

A DNA PROBE IS A SHORT SINGLE-STRANDED SEQUENCE OF DNA WHOSE BASES ARE COMPLEMENTARY TO A SPECIFIC DNA SEQUENCE. THEREFORE IF THE DNA SEQUENCE IS PRESENT, THE DNA PROBE WILL BIND TO IT BY COMPLEMENTARY BASE PAIRING

Question 57

why do we use gene probes

Answer 57

https://merchanttaylorsschools-my.sharepoint.com/:w:/g/personal/tofoma_merchanttaylors_com/Ee8JoqPa3DhHg28xWBu4sToBlqgElLWjLCewcDKnW6eE2A?e=RyFkwi

Question 58

How do gene probes work to detect specific alleles in DNA samples?

Answer 58

Sequence the allele and produce the probe using a gene machine, then amplify using PCR. Label the probe with a fluorescent dye (detected under UV light) or a radioactive isotope (e.g., ¹⁵N). Bind the DNA sample to the bottom of a well (e.g., using a DNA microarray). Incubate with the labelled probe. If the target sequence is present, the probe hybridizes (binds) to it. Rinse the well to remove unbound probes. If the target allele is present, the labelled probe will remain and can be detected.