topic 8- control of gene expression Flashcards
what are the two types of genetic mutations
gene mutations + chromosome mutations
define gene mutation
any change to the quantity of bases or base sequence in the DNA
what is a point mutation
mutation just affects one nucleotide / base e.g substitution
what is a frameshift
base deletion causes subsequent bases to shift left and the whole amino acid sequence to change
a mutation can lead to a non functional enzyme, explain how (6)
change in base sequence of DNA
change in amino acid sequence of enzyme
change in hydrogen / ionic bonds
change in tertiary structure
change in shape of active site
substrate no longer complementary to enzyme
what is cell differentiation
a process by which a cell becomes specialised to carry out a particular function
what is a totipotent stem cell
can differentiate into ANY type of cell (eg. embryonic stem cells - because all body cells develop from these)
why do cells look different and perform different functions
because only the genes that are expressed are transcribed and translated into proteins
so the cell only contains the proteins it needs
can specialised cells specialise into another cell type
no as cell differentiation is irreversible
define stem cell
undifferentiated dividing cells (self renewing)
4 sources of stem cells in mammals
-embryonic stem cells
-umbilical cord blood stem cells
-placental stem cells
-adult stem cells (body tissue of fetus)
locations of adult stem cells
brain, heart, liver
epidermis, bone marrow, adipose tissue, skeletal muscle
what is a pluripotent stem cell
slightly more specialised cells but can differentiate into ALMOST any cell type, apart from placenta (also found in embryos and fetus)
what is a multipotent stem cell
can differentiate into a limited number of specialised cells (eg. stem cells in the bone marrow can produce any type of blood cell but ONLY blood cells) - found in adults (examples - adult stem cells and umbilical cord blood stem cells)
what is a unipotent stem cell
a cell that has committed and can only differentiate into a single cell type (found in adult tissue)
how can a unipotent stem cell be made into a pluripotent stem cell (induced pluripotent stem cell)
unipotent cell is taken from a patient
genetically altered in a lab, various genes are turned on/off to make it similar to the embryonic stem cells
uses of IPS
skin grafts for tissue damage
heart muscle cells for heart damage
B cells of pancreas for type 1 diabetes
skin cells for burns/wounds
describe control of transcription factors by hormones e.g oestrogen
-oestrogen is lipid soluble so diffuses into cell through cell surface membrane
-one in cytoplasm it binds to a receptor on the surface of the transcription factor
-this causes a shape change in the DNA binding site on the transcription factor which activates it
-it enters the nucleus through a nuclear pore and binds to DNA in the promoter region
-this triggers transcription of a particular gene due to RNA polymerase binding
define epigenetics
a study of changes in organisms’ genetics caused by modification of gene expression rather than alteration of the genetic code itself
what is a nucleosome
4 pairs of histones in the core with DNA wrapped twice around the core, another histone holds nucleosome together to stabilise it
2 tags that affect the shape of the DNA histone complex
acetyl groups on histone
methyl groups on DNA
what is the epigenome
the chemical changes to the DNA and histone proteins (chromatin) of an organism - they can be passed down to an organism’s offspring with the DNA
what is acetylation
the addition of an acetyl group to a histone protein donated by acetyl coenzyme A
how does acetylation change the shape of the DNA histone complex
positively charged histone protein tails lose their positive charge
less association between histone and negative phosphate groups on DNA
DNA histone complex does not pack as tightly
what base is methyl often added to in methylation
cytosine
what are the 2 ways methylation of DNA work
methyl groups attract proteins that cause deacetylation of histones
block promoter regions and prevent transcription factors from binding
what can cause biochemical alterations of DNA
smoking , diet changes
describe how translation is inhibited by siRNA
- double stranded RNA is broken up by an enzyme into small interfering RNA
- one of two strands of siRNA binds to enzyme
- the siRNA strand binds to complementary bases on MRNA strand
- the enzyme cuts the mRNA into smaller sections
how can diseases caused by epigenetics be treated
drug to prevent acetylation/methylation
what are chromatin types for tight / loose DNA histone complex
tight - heterochromatin
loose - euchromatin
what is cancer
a group of diseases caused by mutations to genes that control the cell cycle and mitosis thus resulting in uncontrolled growth of cells
how can mutation of protoonco genes cause cancer
only switched on when the cell is going to divide - if mutates to oncogene - becomes permanently switched on - uncontrolled cell division
how can mutation of tumor suppressor genes cause cancer
slow down cell division - gives the cell time to repair mistakes - helps to maintain normal rates of cell division - if mutated cell division speeds up - becomes uncontrolled
what are the two types of tumour
benign - non cancerous
malignant - cancerous
features of benign tumours
-can grow to large size but very slowly
-produce adhesion molecules on cell surface membrane so cells stick together and don’t spread elsewhere
-surrounded by dense capsule - compact
-disrupt functioning of organ
-usually removed by surgery
features of malignant tumours
-grows large and rapidly
-have large nucleus which appears darker due to more DNA
-cells often unspecialised
-do not produce adhesion molecules
-not surrounded by capsule so grow finger like projections into surrounding tissue
-has effects on whole body
-radiotherapy/chemotherapy to treat it
define metastasis
the process by which primary tumours spread resulting in secondary tumours in other parts of the body (through lymphatic/blood system)
how do tumour suppressor genes slow down cell division
-gives cell time to repair mistakes in DNA during replication
-cell to self destruct if it is mutated
what is a mutagenic agent
something that causes increased frequency of mutations/makes you more likely to have a mutation
what is the genome
full set of DNA in a cell/organism
what is sequencing a genome
involves working out the entire DNA base sequence for a
cell/organism
what is the human microbiome project and why is it easier to study prokaryotic cells
the genomes of thousands of prokaryotic organisms are
currently being sequenced (that live on or in human bodies)
the information gained could help to cure many diseases
determining the genome/proteome for prokaryotic organisms is easier because they only have one, circular piece of DNA and have no introns
what is the proteome
full variety of different proteins that a cell/organism is able to produce
why is it more difficult to predict how the genome will be translated into the proteome for 2 reasons
-non-coding DNA (introns)
-regulatory DNA (promoter regions) these can be used to identify the start of a gene
what is recombinant DNA technology
a process by which genes can be manipulated, altered and transferred from one organism to another
steps to in vivo cloning of desired gene
isolation - DNA fragment that contains desired gene is isolated from DNA of human cell
insertion - gene of interest is inserted into a vector (usually bacterial plasmid)
transformation - DNA transferred into suitable host, bacteria that take up plasmid are ‘transformed’
identification - identify transformed bacteria
growth/cloning - bacteria produce large quantities of protein
what are the 3 different ways of isolating a desired gene
-using reverse transcriptase to convert the mRNA of the gene of interest into cDNA
-using restriction endonucleases to cut out the fragment of DNA containing the desired gene from the DNA
-creating the gene in a gene machine (based on its known protein structure)
how to use reverse transcriptase to convert mRNA of desired gene to cDNA
-select a cell that rapidly naturally produces the protein of interest e.g beta cells
-mRNA extracted from cells
-reverse transcriptase uses mRNA as a template to form a single strand of cDNA by CBP
-cDNA isolated by hydrolysis of mRNA with enzyme
-double stranded DNA forms using DNA polymerase
how to use restriction endonucleases to cut out the fragment of DNA containing the desired gene from the DNA
restriction endonucleases are enzymes that cut double stranded DNA by breaking phosphodiester bonds in the backbone, each one cuts at target site
recognition sequences are palindromic
sticky ends - ends exposed to be hydrogen bonded with
how to create a gene using a gene machine
-the amino acid sequence of the protein is determined
-from this the mRNA codons are looked up using a conversion table
-from this the complementary DNA triplets are worked out
advantages of creating genes in gene machine
no introns so can be transcribed by prokaryotes
how to cut vector (plasmid) for insertion of gene
cut with the SAME restriction endonuclease enzyme as the DNA that the gene is isolated from so the sticky ends are complementary
what is a transgenic cell
the organism’s genome now contains foreign DNA/gene (bacteria contains plasmid containing the gene of interest)
how can antibiotic resistance bacteria be used to identify if cells have transformed/transgenic
some cells will no longer be resistant to an antibiotic if vector was cut and gene has been placed in between it
steps to using antibiotic resistant genes
-grow bacteria on agar plate
-transfer bacteria using nylon sheet to plate containing ampicillin antibiotic
-do same with tetracycline
a marker other than antibiotic resistant ones which can determine transformed cells
fluorescent markers
enzyme markers
what is required for polymerase chain reaction
-target DNA
-free DNA nucleotides
-DNA primers (short single strands of DNA)
-TAQ polymerase
-buffer
-thermal cycler
describe 3 stages of PCR
1) temperature of the reaction mixture is increased to 95C degrees, causing the strands of the DNA fragment to separate due to the hydrogen bonds breaking (the heat is denaturing the DNA)
2) mixture is cooled to 55 degrees causing the primers to anneal to their complementary sequences at the ends of the DNA fragment - this provides a starting point for polymerase because polymerase can only add nucleotides onto an existing chain and primers also stop two chains reannealing
3) temperature is increased back to 72 degrees (optimum temp for TAQ polymerase) - TAQ polymerase adds nucleotides onto the primers along each of the strands until it reaches the end - resulting in 2 double stranded copies of the target DNA
what is an example of using PCR
forensic examination - a minute sample of DNA in a single hair or spec of blood - the DNA can be amplified by PCR to analyse
what is a DNA probe
a short, single-stranded length of DNA that has some sort of label attached to it that makes it easily identifiable to us by eye
what are 2 commonly used DNA probes
radioactively labelled probes - made of nucleotides containing phosphorus isotope, can be identified using x ray film
fluorescently labelled probes - emit light under certain conditions (eg. when it successfully binds to the particular target sequence) which can be viewed under UV light
steps of using DNA probe to locate a gene
-DNA probe is made so it has a base sequence that is complementary to gene of interest
(genetic libraries store base sequence for gene)
-double stranded DNA from patient that contains gene of interest is heated to separate strands
-separate DNA strands mixed with probe and cooled
-probe can then bind to its complementary sequence on one of the strands by annealing if the gene is present
-if the probe binds it can be identified by radioactivity or fluorescence emitted by the probe
why are unbound probes washed away
to make sure false positives are not given
why are DNA probes used in genetic screening
to see if individuals have a mutant allele which may give them predisposition to certain diseases, personalised medicine can be provided by doctor to prevent disease
what is genetic fingerprinting
analysis of DNA samples from samples of body tissues or fluids, usually used to identify individuals
what is stage 1 of genetic fingerprinting
extraction
-DNA is extracted from individuals sample (tissue/fluid) by separating DNA from rest of the cell
-can be increased using PCR
what is stage 2 of genetic fingerprinting
digestion
restriction endonuclease enzymes are used to cut up the DNA from the sample into fragments
what is stage 3 of genetic fingerprinting
separation
-mixture of DNA fragments are placed into a well in agarose gel and a voltage is applied
-DNA is overall negatively charged due to the oxygen in the phosphate groups
-the fragments are attracted to and move through the gel towards the positive electrode
why do the DNA fragments move different distances in the agar
the larger fragments travel more slowly through the gel (more resistance) and smaller fragments travel quicker and further in a certain amount of time
what do you add to the well to compare DNA fragments to
DNA fragments of KNOWN sizes
what is stage 4 of genetic fingerprinting
hybridisation
- DNA sequence of interest can be found by adding a fluorescent or radioactive labelled probe that is complementary
-before the probe is added - alkali is added to separate the double stranded DNA
-any excess probe is then washed off
why can’t the probe be added to the gel
would get caught up in the gel and gel would dry up
what is stage 5 of genetic fingerprinting
development
probes position in gel can be determined
advantages of in vitro cloning
-it is extremely rapid
-does not require living cells, only base sequence of DNA
advantages of in vivo cloning
-particularly useful to introduce gene into organism
-almost no risk of contamination
-very accurate with no errors
-it cuts out specific genes
benefits of recombinant gene technology
-microorganisms can be modified to produce a range of substances that can be used to treat disease
-replacing defective genes can be used to cure certain genetic disorders
-genetically modified animals can produce expensive drugs cheaply
risks of recombinant gene technology
-engineered gene mutating could turn organisms into pathogens
-any manipulation of the DNA within a cell will have
consequences for the metabolic pathways of that cell
-antibiotic resistant bacteria used in in vivo cloning could spread antibiotic resistance to harmful bacteria
