Control of Gene Expression Flashcards
Define Mutation
Mutations are any change to the nucleotide base. They can be caused by errors during DNA replication.
State and Define the 6 types of mutation.
Substitution- One or more bases are swapped for another
Deletion- One or more bases is removed
Addition- One or more bases are added
Duplication- One or more bases are repeated
Inversion- A sequence of bases is reversed
Translocation- A sequence of bases us moved from one location to the genome of another.
Why can mutations lead to problems?
The order of DNA bases in a gene determines the sequence of amino acids in a particular polypeptide, if a mutation occurs, the sequence of amino acids could change subsequently change the tertiary structure of a protein changing its function.
Why can mutations have no negative outcomes?
Not all mutations cause a change in the protein. The degenerate nature of all genetic code means some amino acids are coded by more than one DNA triplet.
What is a frameshift?
Frameshift- When the mutation causes a change in the number of bases so the triplet code is read in another way.
What is a mutagenic agent and give examples?
Physical or chemical agents that increase the rate of mutations
Examples include UV and ionization radiation as well as some chemicals and viruses.
What are the three ways mutagenic agents could increase the rate of mutations?
Act as a Base- Substituted for a base during replication
Altering Bases- Some chemicals delete or alter bases
Changing DNA structure- Some types of radiation can change the structure of DNA causing replication problems
How do tumours form and how do they become cancerous?
When mutations occur in genes that control the rate of cell replication, it can cause uncontrolled cell division forming a tumour.
Tumours that invade and destroy tissues are called cancers.
State and define the two types of tumours.
Malignant Tumour- These are cancerous. They usually grow rapidly and invade all tissues. Cells can break off and spread to other parts of the body in the bloodstream.
Benign Tumour- Not cancerous, they usually grow slower than malignant tumours and are covered in fibrous tissues that stops cells invading. They are usually harmless but can cause blockages and preassure.
What are the two types of genes that control cell division?
- Tumour Suppressor Genes
- Proto-oncogenes
Talk about Tumour Suppressor Genes and how they can cause a tumour.
When functioning normally, these slow cell division by producing proteins that stop dividing or cause them to self- destruct.
If a mutation happens in this gene, the gene will be inactivated and the protein it codes for isn’t produced so the cell divides uncontrollably
Talk about Proto-Oncogenes and how they can cause a tumour.
When functioning normally, they stimulate cell division by producing proteins.
If mutation happens, the gene can become overreactive which stimulates the cells to divide uncontrollably.
What are Oncogenes?
these are formed from mutated proto-oncogenes and result are permanently switched on resulting in cell division that is uncontrolled.
It does this by permanently activating a cell surface receptor or coding for a growth factor.
What is Methylation and why is it important in gene expression?
Methylation is the addition of methyl onto something. Methylation of DNA is an important method of regulating gene expression as it can control whether or not a gene is transcribed and translated.
- at CpG site, with no change to DNA base sequence
- chromatin more tightly packed (heterochromatin)
- harder for RNA polymerase to bind
Talk about abnormal methylation of tumour suppressor genes.
HYPERMETHYLATION- too much
Genes are not transcribed so the proteins they produce to slow cell division aren’t made. This means. that cells are able to divide uncontrollably causing tumours
HYPOMETHYLATION- too little
The hypomethylation of Proto-Oncogenes causes them to act as oncogenes, increasing the production of the proteins that encourage cell division, which could lead to tumours
Talk about the role of Oestrogen in Brest Cancer.
Some women may be exposed to more oestrogen than others due to starting menstruation earlier, or the menopause later, which could overextended period of time increase the risk of breast cancer.
This could be due to oestrogen stimulating certain breast cells to divide and replicate or some research suggests that oestrogen is able to introduce mutations directly into the DNA of certain breast cells.
What are the two categories of risk factors for cancer?
Genetic Factors- Cancer can be linked with specific inherited alleles If you inherit that allele, you’re more likely to get that type of cancer
Environmental Factors- Exposure to radiation lifestyle choices such as smoking, increased alcohol consumption, and a high fat diet have all been linked to an increased chance of developing some cancers
What is a stem cell?
Stem cells are undifferentiated cells which can keep dividing to give rise to other types of cells
State and define each type of stem cell.
Totipotent- cells in the embryo which can form any type of cells in the body and embryonic cells
Pluripotent- can form any cell in the body but not embryonic cells. They are also found in early embryos and can help replace damaged tissues.
Multipotent- can differentiate into other cells but are limited i.e. bone marrow
Unipotent- can only differentiate into one type of cell
What are induced pluripotent stem cells?
Pluripotent stem cells created from unipotent stem cells.
Give an example of a stem cell therapy.
Bone Marrow Transplant:
Bone marrow transplants can be used to replace the faulty bone marrow in patients that produce abnormal blood cells. The stem cells in the transplanted bone marrow divide and specialise to produce healthy blood cells. This has been used successfully to treat some genetic disorders such as sickle cell anemia.
How are stem cells obtained?
Adult Stem Cells- Obtained from the body tissues of an adult and the bone marrow
Embryonic Stem Cells- IVF, When an egg cell is fertilised by sperm outside the body. in early stages of development, stem cells are removed and the rest of the embryo is destroyed.
Talk about ethical considerations of Stem Cells?
Some people believe that at the moment of fertilisation, an individual is formed that has the right to live so they believe it’s wrong to destroy the embryos
While other people believe that because stem cells are obtained from egg cells that haven’t been fertilised by sperm, but have been artificially activated to start dividing, that they can be used.
As adults are able to consent to stem cell removal. And it results in little injury and no death. People believe we should only use adult stem cells
However, because adult stem cells aren’t as flexible as embryonic stem cells, they have less use in scientific research
What are transcription factors and how do they work?
These are proteins that bind to specific sites on a DNA sequence, determining transcription
They bind to promotor regions near the start of the gene allowing them to control the rate of transcription and subsequently the level of gene expression.
What are the two types of transcription factor?
Activators, which increase the rate of transcription, helping RNA polymerase binds to the promoter and start transcription
Represses, which reduce the rate of transcription by blocking the binding of RNA polymerase.
Talk about oestrogen in the regulation of transcription and translation
The Hormone oestrogen has the ability to alter transcription through altering Transcription factors:
1. As oestrogen is lipid soluble, it can freely diffuse across the membrane and bind to a receptor molecule on a transcription factor
- The finding alters the shape of theDNA binding site on the transcription factor, making it able to bind to the DNA
- The transcription factor therefore enters the nucleus via the nuclear pore, where it binds to DNA. This stimulates the transcription of the gene that makes up the DNA
What are the two types of RNA repressors?
Small Interfering RNA (siRNA)- Fully complementary to target, meaning they can only bind to a specific mRNA sequence
MicroRNA (miRNA)- Partially complementary to target, meaning it combined to different parts.
Talk about siRNA in the control of transcription and translation.
It is used for short term switching off of genes. The siRNA binds to a complementary sequence of mRNA, As mRNA is usually single stranded. The cell therefore detects the double strand on mRNA and views it as abnormal. This causes the mRNA to be broken down by Enzymes preventing translation
Talk about miRNA in the control of transcription and translation.
miRNA is first processed into two single strands. One strand associates with proteins and binds to target mRNA Which essentially blocks the translation of the target mRNA.
What is Epigenetics?
The study of heritable changes to the phenotype that don’t involve changes to base sequences. This means it doesn’t involve mutations
What is an epigenome?
The chemical compounds and proteins that can attach to DNA and histones, affecting how genes are expressed.
What are the mechanisms of epigenetic control?
- DNA methylation
- Acetylation of Histones
Talk about DNA methylation in epigenetic control.
A methyl group is added to the DNA sequence at a CpG site inhibiting the transcription by physically blocking it or attracting proteins that condensed chromatin making it more tightly packed
This prevents transcription factors and RNA polymerase from binding to the promoter region silencing the gene
Talk about Acetylation of histones in epigenetic control
When histones are acetylated, the chromatin is less condensed. This means that the transcriptional machinery can access the DNA allowing genes to be transcribed.
When Acetyl groups are removed from the Histones, the chromatin becomes highly condensed, and genes in the DNA can’t be transcribed because transcription factors can’t physically access them.
Talk about epigenetics and cancer
Epigenetics can increase the risk of developing cancer increase. methylation of tumour suppressor genes reduce their expression, which can lead to cancerous tumours.
Heterochromatin
tightly packed, harder for transcriptional enzymes to access DNA so gene is switched off
(DNA methylation)
Euchromatin
loosely packed, easier for transcriptional enzymes to access DNA so gene is switched on
(acetylation)
Talk about DNA methylation inhibitors
Drugs that prevent methylation of tumor supressor genes, keeping them active to slow cell division and prevent tumours
Since epigenetic changes occur in many cells, treatments must be highly specific, for example, cancer drugs often target only dividing cells to minimize damage to normal tissues
What is a genome project?
A project of mapping the entire DNA nucleotide base sequence of a range of organisms.
The idea is to map the DNA base sequences that make up the genes of the organism and then map these genes on the chromosome. This makes a complete map of all genetic material (genome)
Talk about sequencing genomes in simple organisms
The genome structure is relatively straightforward as they have no or little non- coding regions- so there’s a more direct correlation between the genome and proteome
Talk about applications of sequencing simple organisms DNA.
Vaccine Production: By identifying proteins and potential antigens from pathogens, researchers can pinpoint targets for vaccine development.
Drug Discovery: Knowledge of protein structures and functions aids in the design of new therapeutics, by targeting specific proteins involved in disease processes.
This can lead to better management of the spread of infection
Talk about sequencing DNA of complex organisms
The genome of complex organisms not only contains regions of coding DNA, but also introns- non- coding regions and regulatory elements- promoters, enhancers, silencers and insulators controlling gene expression.
This means the genome doesn’t directly align with the proteome.
Talk about the development of more sequencing methods
Sanger Sequencing- early method, this was time- consuming and expensive
Next- Generation Sequencing (NGS)- this the modern method which is highly automated allowing researchers to process many samples simultaneously with increased efficiency and accuracy.
- Talk about the good implications of genome projects
- provide insight into evolutionary relationships
- help identify genetic variations linked to disease
Challenges of genome projects
- ethical cosiderations, data privacy, consent, genetic discrimination, design babies.
What is recombinant DNA tehnology?
This is a for of genetic engineering involving the transfer of DNA from one organism into another resulting in the organism having recombinant DNA and becoming transgenic organisms
How is recombinant DNA technology able to happen?
The genetic code is universal meaning almost every organism uses the same bases and the same codons to code for the same amino acids
Also, the mechanisms of transcription and translation are also universal
What are the steps for genetic engineering?
- Identification- of the DNA fragment or gene
- Isolation of desired fragment
- Multiplication of DNA fragment
- Transfer into organism using a vector
- Identification of cells with new DNA fragment- using a marker
What are the ways of isolating DNA fragments?
Reverse Transcriptase
Restriction Endonuclease
The gene machine
Talk about reverse transcriptase to isolate genes
mRNA is combined with reverse transcriptase and nucleotides to create a single strand of complementary DNA (cDNA). DNA polymerase can then convert cDNA into DNA.
Talk about restriction endonucleases to extract genes
These bind to specific restriction sites on DNA and cut it. They cut it in either a straight (blunt ends) or uneven (stick ends)
Talk about the gene machine.
This is when scientists use computers to generate nucleotide sequences to produce a gene.
Talk about the process of creating DNA fragments using the gene machine
- identify the target sequence
- Feed the desired sequence into a computer
- The computer designs short, single strands of nucleotides that match the sequence called oligonucleotides
- These are joined together to form a complete gene
- use PCR to amplify
- Insert into a vector
Evaluate the gene machine
- Useful when the gene has to be introduced into an organism
- no risk of contamination as the same restriction endonuclease is used to produce sticky ends.
- Mutations are rare
- cheap
- However, DNA fragments have to be isolated from cell components
- Slow
What are gene markers
These are genes that are incorperated into the plasmid so the cells with the DNA can be seperated. There are different types: antibiotic resistant, flourescent and enzyme.
What’s the difference between fluorescent markers and enzyme markers?
Fluorescent markers- gene from jellyfish to plasmid making a fluorescent protein GFP. A lack of GFP shows the gene hasn’t been incorporated
Enzyme markers- the gene that produces the enzyme lactase is used as a gene marker. bacteria wont produce this gene so wont change any colour of substrate.
What is replica plating?
Replica Plating- This is the use of an antibiotic gene as a marker. When colonies have specific gene markers—for example, a marker that provides antibiotic resistance—their ability or inability to grow on these selective plates reveals which colonies possess the marker. This makes it a valuable tool for screening mutants and studying gene function based on observable traits.
Describe the process of producing DNA fragments
- Restriction endonuclease or reverse transcription is used to produce DNA fragments containing required genes.
- This is inserted into the plasmid of a vector using DNA Ligase
- Include a marker gene in order to identify what cells have the desired genes
- These are introduced to host cells
- Using gene markers, identify the host cells with the DNA
- Culture these cells to produce on a large scale
What are the two amplification methods and what’s the difference?
In vitro- polymerase chain reaction
In vivo- using bacteria
What is needed for PCR?
- DNA fragments - to be copied
- Taq polymerase to join nucleotides to a template strand
- Primers- to start the process
- Nucleotides
- Thermocycler- change the temperature
What are the three steps for PCR?
- Separation of strands- The DNA fragments, primers and Taq polymerase are placed into a vessel in the thermocycler. The temperature is increased to 95 degrees to separate the two strands by the breaking of hydrogen bonds.
- Annealing of the Primers- The mixture is cooled to 55 degrees to join primers to their complementary bases at the end of the DNA fragment. The primers provide the starting sequence for DNA polymerase to being DNA copying. They also stop the strands from joining.
- Synthesis of DNA- the temperature is again increased to 75 degrees. This is the optimum temperature for the DNA polymerase to add complementary nucleotides along the DNA strands. More than 1 million strands are made in 25-cycles
Evaluate the PCR
- Rapid
- Doesnt need living cells
- Only the target gene is copied
- However, only small DNA fragments
- doesnt copy mRNA or proteins, only DNA
- expensive
Talk about the process of in vivo cloning
Insertion:
- DNA cut from genes is transferred to a vector which is usually a plasmid
- Restriction endonucleases cut DNA fragments making sticky ends
- DNA fragments mix with opened plasmids by DNA ligase making recombinant DNA
Transformation:
- Plasmids are then reintroduced to host cells. They have to be:
- Ice cold calcium chloride solution- make more permeable
- Then heated to 42 degrees for 1-2 mins.
- This encourages the cells to take in the plasmids
- However, not all host cells are taken up- use of markers.
What is recombinant DNA technology?
Microorganisms, plants and animals can all be transformed using recombinant DNA technology- genetic engineering. Transformed microorganims can be made using the same technology as in vivo cloning.
Give an example of recombinant DNA tech and the process. (insulin)
foreign DNA can be inserted into a microorganism to produce insulin:
- DNA fragment containing the insulin gene is isolated
- The DNA fragment is inserted into a plasmid vector
- The plasmid containing the recombinant DNA is transferred into a bacterium
- Transformed bacteria are identified and grown
- the insulin produced is extracted and purified.
Talk about transformed plants
a gene that codes for a desirable protein is inserted into a plasmid. This is added to bacteria which is used as a vector to get the gene into the plant cells. They can be transformed so they give a higher yield or are more nutritious to reduce the risk of famine and malnutrition. Crops can also be transformed to have resistance to pests or droughts reducing the costs of any environmental problems.
Talk about transformed animals
a gene that codes for a desirable protein can be inserted into an early animal embryo or into an egg making animals with desirable traits. Again, this can include things like fatter chickens for more meat or animals that require less food using less energy, resources and money.
Talk about recombinant DNA and industry
Enzymes can be produced from transformed organisms so they can be produced in large quantities for less money, reducing costs.
Talk about recombinant DNA in medicine
Many drugs and vaccines can be produced by transfored organisms. Drugs made using this DNA can be produced quickly, cheeply and in large quantities.
Talk about problems of recombinant DNA technology
Monoculture farming may make the whole crop vulnerable to tje ame disease as genetically identical, they also reduce biodiversity ruining the environment. Superweeds are resistant to herbicides- unknown consequences.
Without proper labelling, people may think that they wont have a choice about whether to consume food. May also kick smaller businesses out of business.
Worry it could be used unethically ie, designer babies
What is gene therapy?
This is the altering of defective genes inside cells to treat genetic disorders and cancer. How you do this depends on wheter the disorder is caused by a mutated dominant allele or two recessive ones.
