Molecular biology 2.7, 3.1, 3.2, 3.5, 7.2, 7.3 Flashcards
What is a gene?
- heritable factor
- consists of a length of DNA
- influences a characteristic
What is an allele?
> one specific form of a gene
occupy the same locus in different specimens
differ by one (SNPs) or a few bases
formation of new alleles
- point mutation - change of one or some bases within a gene
- substitution (most common), insertion, deletion, inversion
- silence: no change, missense: different, nonsense: no protein
What is the structure of haemoglobin?
- protein blood (red)
- 4 polypeptide chains
- 2 alpha and 2 beta
- ring-like heme (with Fe)
- oxygen binds to iron
What are the causes of sickle cell anaemia?
- single base substitution
- CTC → CAC
- mRNA: GAG → GUG
- glutamine 6 (in beta chain) → valine
- amino acids
- Hb^A - in healthy humans
- Hb^S
What are the results of haemoglobin mutation (sickle cell anaemia)?
- deoxygenated haemoglobin stick to each other
- stiff fibres in red blood cells
- deformation of RBC
- fragile - from 30 to ~4 days
- inability to replace them causes anaemia
What is a mutation?
- random change
- no mechanism behind it
- most common: base substitution
- one base is exchange for another
What are the steps of mutation and potential causes?
- change of base in gene sequence
- change of codon in RNA sequence
- (potential) change in amino acid sequence
- (potential) change in protein structure
- (potential) change of loss of protein function
- change of organism’s physiology
- mutations are random = never beneficial
- neutral or harmful
- can be passed (in gametes)
What is a genome?
- whole DNA of an organism
- coding and non-coding parts
- 46 linear chromosomes + DNA in mitochondrion (in humans)
- in plants: chromosomes + DNA in mitochondrion and chloroplasts
- in prokaryotes: circular DNA
What is the link between malaria and sickle cell anaemia?
> malaria
- different red blood cells shapes prevents from reproduction of Endemic falciparum (malaria protist)
What was the Human Genome Project?
- base sequence of a whole human genome
- mine of data
- which sequences code for proteins
- non-coding parts of DNA are important
- comparison between humans and other species
How do chromosomes differ in eukaryotic and prokaryotic cell?
- eukaryotes
- linear (in nucleus)
- forms nucleosomes
- non-coding sequences
- linear (in nucleus)
- prokaryotes
- circular
- nucleoid = DNA + cell membrane
- compact (no repetitions)
What is a chromosome?
- entire chain of DNA along with a group of stabilising proteins
What was the procedure of Cairn’s technique and what did the results show?
- autoradiography
- DNA labelled with radioactive thymidine
- during DNA replication
- isotope incorporated into new DNA strand
- the cell was placed in dialysis membrane and lysozyme was used to break down its walls
- DNA in membrane
- radioactivity allows exposure of photosensitive film (radioactively-sensitive emulsion)
- tritium decayed
- image on film
- autoradiograph
- conformation of ‘replication fork theory’ → bidirectional
- estimation of chromosomal / DNA length
- tritium decayed
What are plasmids and what is their role?
- in prokaryotes
- small circular DNA bits (naked)
- exchanged between bacteria
- antibiotic resistance
- plasmids do not replicate in the same time as DNA
- not passed during division
What is the role of nucleosome?
- protecting DNA from harm
- changing structure
- information in DNA used or not
What is the structure of nucleosome?
- 8 histone proteins (+ charge)
- histone tails reach out to neighbouring nucleosomes
- tightly packing
- histone tails reach out to neighbouring nucleosomes
- 2 loops of DNA (- charge phosphate group)
- differences in charges cause DNA to wrap around histones
- in nucleus
- enzymes modify tails
- weakening interactions between nucleosomes
What are homologous chromosomes?
- pair
- one from each parent
- corresponding sequences
- during meiosis they pair
- 46 chromosomes
- 2 sets of 23 chromosomes
How many chromosomes are in human cell?
- haploid - 1 chromosome of each kind: 23
- sex cells (to reproduce)
- diploid - 2 homologous chromosomes of each kind: 46
- polyploid
What is a karyogram?
- homologous pairs of chromosomes of decreasing length
What are sex chromosomes?
> XX - female > XY - male - Y is smaller > human body functions without duplicate of X (in XY) - for XX - X-inactivation (random in each cell) - differences between twins
Why does genome size vary and what does it depend on?
> size and number of chromosomes
complexity (not proportional)
some organisms may have many non-coding areas
What are promoters?
> binding site of RNA polymerase
- determine whether transcription occurs
What are expression regulating sequences?
- enhancer
- silencer
- promoter-proximal sequence
- terminator - ending part
What is an operon?
- group of proteins having similar functions
- sequenced together
How is gene expression of absorption of lactose regulated in E. coli?
- gene expression is dependent on environment
- operon: lacZ, lacY, lacA
- break down lactose (glucose for energy)
- repressor - protein inhibiting expression of breaking down lactose (if there’s no lactose in cell)
- RNA polymerase cannot bind to operator (promoter)
- when lactose is present, it binds to repressor
- expression of gene starts
- presence of glucose
- bacteria uses glucose (easier)
- CAP site
- CAP protein binds (low glucose)
- increased expression
- cAMP helps with binding
- high level of transcription
- CAP protein binds (low glucose)
- high glucose levels
- no cAMP produced
What happens during cell division?
> cell may divide into 2 daughter cells
- different functions = different expression rates
What is the direction of transcription?
- RNA strand: 5’ → 3’
- sense strand is the same
- antisense strand: 3’ → 5’
How does environment impact gene expression?
- amount of sunlight
- temperature
- Siamese cats
- mutant allows for production of pigment only in low temp.
- Siamese cats
What happens in the first step of transcription (initiation)?
- promoter (usually TATA)
- transcription factors bind to DNA - initiation complex
- find promoters
- help binding of RNA polymerase
- ATP is added to start the process
- ADP and P
- released when transcription begins
- activators and repressors (increase and decrease the rate)
- silencers and enhancers
- eukaryote: set of proteins
What happens in elongation (transcription)?
- as RNA polymerase goes it unwinds the helix
- RNA polymerase transcribes the DNA sequence
- producing RNA strand by attaching complementary base pairs
What happens in termination (transcription)?
- transcription stops as the terminator sequence is reached
- RNA dissociates from RNA polymerase
- pre-mRNA is formed
What are the post-transcription modifications (capping and poly-adenylation)?
- only in eukaryotes
- preparing RNA for translation
- quality control
- protection from damage
- capping
- addition of 5’ cap
- protects the end
- poly-adenylation
- adding A bases (3’)
What happens during mRNA splicing and what is its role?
- introns (non-coding) are cut out
- later used
- jointing exons
- alternative splicing
- some exons can be cut out
- make products more complex (different variations)
- made by spliceosomes
What is epigenetics?
- chemical modifications of DNA or proteins affecting DNA
- during development
- reset when forming gametes
How is transcription regulated (changes in DNA and chromosomes)?
- DNA methylation
- direct modification
- adding ethyl group to cytosine
- interactions of proteins with DNA
- gene silencing
- modification of histones
- histone tails have positive charge
- acetylation = adding acetyl group
- neutralisation of histones
- DNA is negative
- less coiling
- more transcription
- methylation = adding methyl group
- DNA more coiled (condensation)
- less transcription
What happens to epigenetic tags in gametes?
- dividing cells still have tags
- when joined together cells tags are erased
- cells are reprogrammed
- parts of the tags can stay
What happens during translation?
> polypeptides are synthesised
> amino acid sequence determined by base sequence of mRNA
Where does translation take place?
- in cytoplasm (rough endoplasmic reticulum)
- ribosomes
What is the structure of ribosome?
- 2 subunits: large and small
- rRNA
- binding site for tRNA
- 2 tRNAs can bind at the same time
- A (aminoacyl) site
- recognition and interaction of codon and anticodon
- P (peptidyl) site
- holds peptide-caring tRNA
- E (exit) site
- release discharged tRNA
- binding site for mRNA
- unit ‘S’ (svedbergs)
- measure of speed of molecule in centrifuge
- do not add up
- once bound subunits lose their mass
- prokarya: 70S (50S big, 30S small)
- eukarya: 80S (60S big, 40S small)
What are codons?
- sequence of three bases of mRNA that are translated into 1 amino acid
- START codon - Methionine
- STOP codon inhibit translation
- 64 possible codons (4^3)
- 24 amino acids
- some codons code for the same aa
- genetic code is degenerate
- some codons code for the same aa
Why is genetic code universal?
- each codon translates into the same amino acids among different organisms
- there are some exceptions: mitochondria, yeast, protists
What is the role of mRNA in translation?
- produced in transcription
- information about sequence
- structure
- reading part (from START to STOP)
- 5’ → 3’
What is the structure of tRNA?
> anticodon
- complementary sequence that reads codons
site for attaching amino acid
- sequence CCA (3’ end)
How is tRNA activated?
- tRNA-activating enzyme (aminoacyl-tRNA-synthetase)
- active site of enzyme is anticodon and amino acid specific
- ATP = AMP + PPi
- ester bonds between tRNA and amino acids
- process
- amino acid and ATP binds
- amino acid activation by AMP
- pyrophosphate is released
- tRNA binds to enzyme
- amino acid bind to tRNA (covalent bond)
- AMP released
- charged tRNA released
What happens in initiation of translation?
- positions small subunit (near START of mRNA)
- tRNA with Met binds
- in P site
- big ribosomal unit attracted
- elongation starts
What happens in elongation of translation?
- peptidyl-tRNA binds to P site
- charged aminoacyl-tRNA enters A site
- anticodon binds to codon
- peptide transferase transfers polypeptide to A-bound tRNA
- translocation
- ribosome moves 3 nucleotides ahead towards 3’ end
- discharged tRNA → E site
- peptide-carrying tRNA → P site
- A site emptied
- discharged tRNA → E site
- ribosome moves 3 nucleotides ahead towards 3’ end
What happens in termination of translation?
- STOP codon is reached
- tRNA is released
- polypeptide dissociates
- ribosomal units separate
How does translation occur in prokaryotes?
> transcription and translation occur simultaneously
- no nucleus
- no introns
- less ability to control the process
polysome
- multiple ribosomes translating the same mRNA
What is the difference between bound and free ribosomes?
> free ribosomes - produce proteins for usage in cell > bound ribosome - on rough endoplasmic reticulum - proteins for secretion
What are different protein structures?
- primary structure
- covalent peptide bonds
- sequences of polypeptides
- secondary structure
- H-bonds between carboxyl group (C=O) and amino group (N—H)
- beta-pleated sheets
- alpha-helix
- tertiary structure
- 3D structure
- R group with water medium
- R group + with R group -
- hydrophobic amino acids stick to each other
- polar R-groups form H-bonds
- disulphide bridges
- quartenary structure
- more than 1 polypeptide
- non-polypeptide elements
How does gel electrophoresis work?
- separate proteins according to size
- process
- DNA samples are put in a gel
- charged negatively so it will move towards + side
- small fragments move faster
- DNA samples are put in a gel
What is the role of DNA profiling?
- paternity tests
- DNA of mother, child and potential father
- forensic investigations
- blood stains, hair, semen
- DNA of victim and suspects are compared with the sample
What are stages of DNA profiling?
> sample DNA obtained
sequences of DNA that vary are copied by PCR
- split into fragments (restriction endonucleases)
gel electrophoresis
pattern of bands obtained
- compared with suspect and victim
How does genetic modification work?
- genetic code is universal
- genes transferred between species
- new characteristics
- genetically modified organism (GMO)
- DNA from other species (US definition)
- crossing is not GMO
- transgenic organisms
How are genes transferred to bacteria?
> mRNA secreted from human cells
- reverse transcriptase
- complimentary DNA (cDNA)
from bacterium cell plasmid is obtained
- part of it cut with restriction enzyme
- sticky ends = cutting 2 strands at different points
cDNA and plasmid fuse
- DNA ligase
new plasmid introduced to bacterium
bacterium replicates
- gene expression of new fragment (ex.: insulin)
bacteria is separated and substance purified
available for usage
Assessment of one benefit and risk of GM crops
- example: Golden Rice
- rice that produces beta carotene
- supply of vitamin A needed in countries where a lot of rice is consumed (Asia)
- benefit (health): the nutritional value of crops is improved
- risk (environmental): cross-breeding (or spreading) to wild plants, uncontrollable effects
- contaminating wild rice
Assessment of risks and benefits of GM of Bt corn
> plants produce toxins that kill insects
benefit (environmental): less pesticides are used
risk (environmental): non-target organisms could be affected by toxins
- monarch butterfly
- feed on milkweed near corn
What are clones?
- genetically identical organisms
- single parental cell
- asexual reproduction
- twins (monozygotic)
- zygote divides into 2 cells
- 2 embryos
What are examples of natural cloning?
- garlic bulb
- by photosynthesis enough energy to produce group of bulbs
- strawberry
- plantlets at the end of plant
- grow roots
- independent from parent plant
- plantlets at the end of plant
How can animals be cloned?
> embryos are pluripotent
- can develop in every tissue
embryo broken artificially can produce clones
characteristics cannot be chosen
How can adult animals be cloned?
- cells are differentiated
- in frog
- once a nucleus is put in a new cell it develops as if a zygote
- in humans — stem cells
Producing Dolly
- somatic-cell nuclear transfer
- cells taken from udder of adult sheep
- cultured in lab
- unfertilised egg from another sheep
- nucleus removed
- egg cell formed
- pulse of electricity
- embryo transferred to uterus of another sheep
- surrogate mother gave birth to Dolly
