Genetics S1 Y1 Flashcards
How is the bacterial genome replicated?
Plasmid replication - replication begins at the origin of replication and the strands separate and replicate until two daughter plasmids separate (made up of a new and old strand)
2 characteristics of plasmids that make them useful
- Used for adaptation (not essential for life though.
- Promote genetic exchange or encode genes to kill other bacteria
What enzyme is used to break the double helix structure of DNA?
Why is it used?
-Topoisomerase ii
-Used to supercoil the DNA to reduce the space it takes up
How is the genome structured in eukaryotes?
How was this discovered?
-In nucleus, arranged into linear chromosomes that are comprised of chromatin condensed with histones. The chromosomes have many origins of replication.
-Giemsa patterning
Levels of structure to form a chromatid?
DNA associated with histones (condense + support) to form nucleosomes which fold to form a chromatin fibre which is coiled and condensed to form a chromatid
Why are mitochondria and chloroplasts different to other organelles?
-They were both engulfed via endocytosis and were eubacteria before they became associated with eucarya.
-They exchanged genetic material with the nucleus - now depend on nuclear transcription e.g. the proteins in mitochondria are encoded in the nucleus and must be translated and taken up by the mitochondria
-They have much smaller genomes as most of their genome has migrated to the host nuclei
-What were chloroplasts and mitochondria before they they became symbionts (closely evolved association via endosymbiosis)?
-Which parent are they inherited from?
-Chloroplasts = cyanobacteria
Mitochondria = proteobacteria
- Female parent
What are the genes in mitochondria for? (4)
- Respiration and oxidative phosphorylation
- Transcription and translation
- RNA processing
- Importing proteins into cell
Why is the mitochondrial genome shorter than the normal genome?
Contains fewer genes, has no introns, only one promoter per strand, 2 strands
Why are diseases associated with mitochondrial mutations complex?
Mitochondria segregate randomly during cell division, so random daughter cells will contain mutated mitochondria. Even if a mother shows no signs of mitochondrial disease, she may produce diseased gametes that means her offspring may get symptoms of mitochondrial disease
What is variation in mitochondrial DNA called?
Heteroplasmy
How is it possible that embryonic stem cells with the same genome can be differentiated to form many types of cell?
Differential gene expression (different genes are expressed)
What is the promoter region used for?
To initiate transcription (where transcription factors and relevant proteins such as DNA pol. bind)
6 stages of gene expression regulation?
Chromatin –> transcription –> RNA processing –> RNA stability –> translation –> posttranslational modification
- Why must chromatin condensation vary?
- How does it vary?
- Must be less condense when transcription is occurring so proteins can access it
- Histone tail methylation and acetylation, DNA methylation
What does DNA methylation cause?
Changes chromatin structure, nucleosome positioning and modifies histones
What is the effect of heavy DNA methylation (especially on CpG islands)?
It prevents transcription (and silences corresponding genes when CpG islands are methylated)
How is the issue of 2 copies of the X chromosome dealt with in females?
One X chromosome is inactivated through engulfing by RNA to change the chromatin structure
How are transcription factors involved in transcription?
Two main types are repressors and activators:
- Repressors bind to promoter region and inhibit translation
-Activators bind to enhancer regions and increase rate of transcription
How do all genomes in all organisms relate?
All have ROUGHLY the same number of exons (individual of their size)
What organism has the largest genome?
Protozoa
What is the C-value paradox?
Term given for the lack of a link between an organism’s complexity and the size of its genome
2 reasons why eukaryotic genomes vary in size?
- Polyploid chromosomes
- Levels of introns vary
2 things repeated sequences in a genome can be?
Dispersed or tandem (clustered)
What are transposable elements?
Sequences that can replicate and insert into new positions - this can restore normal function of a gene but it can reinsert somewhere where another gene’s function is disrupted
What is the difference between negative and positive supercoils?
Negative = underwinding (less tight) - MORE COMMON IN EUKARYOTES
Positive = overwinding (more tight)
What do supercoils in bacterial DNA form?
A nucleoid
Which eukaryotic cells do not have mitochondria and why?
Those that live in anaerobic environments (have hydrogenosomes instead)
2 hypotheses for the origin of eukaryotic cells?
- Evolved from ancestral archaeon that later incorporated the proteobacterium (mitochondria)
- Evolved from symbiosis of archaeon and proteobacterium (mitochondria)
3 ways some genes are regulated by RNA degradation?
- 5’ cap removed (prevents initiation of translation)
- poly (A) tail shortened
- Degradation of 5’ UTR, coding sequence and 3’ UTR
What is the poly (A) tail for and what does it enhance?
- They act as cap-binding proteins and prevent degradation
- They enhance binding of the 5’ end of mRNA to the ribosome
2 ways RNA is modified to make proteins for diverse?
- Alternative splicing (removal of some exons and occasionally introns so one gene can produce many products)
- RNA editing (modifies nucleotides to change protein)
3 ways RNA interference is used to regulate genes?
- Translation inhibition (via small and microRNAs e.g. by methylating DNA or histones or degradation)
- Transcriptional silencing (by changing chromatin)
- Lowering gene expression and degradation by siRNAs
How do siRNAs and miRNAs prevent translation?
Form DNA hairpin and then enzymes cleave the stem to form double-stranded fragments - one of these strands of RNA enters RNA-induced silencing complex (RISC) to result in degradation and inhibited translation
2 types of transcriptional regulation?
- Positive regulation - activator binds to DNA near gene to allow transcription (RNA polymerase can bind to promoter)
- Negative regulation - repressor binds to DNA near gene to inhibit transcription (RNA pol. cannot bind to promoter)
How do activators work?
Bind to enhancer region and cause an allosteric change in DNA so RNA pol. can bind
- How do repressors work?
- How does an inducer affect this process?
- Bind to operator and cause allosteric change so RNA pol. cannot bind
- They allow transcription as they bind to repressors and change their structure so they cannot bind
What is an operon?
Cluster of structural genes and sequences that control transcription that are transcribed together
Lac operon:
- What is produced to break down lactose?
- How is it ensured that the enzyme is not synthesised when lactose is not present?
- How does this change when lactose is present?
- Beta-galactosidase
- Repressor coded for by lacI is bound to operator so lacZ (B-galactosidase) and lacY (lactose permease) cannot be expressed
- Inducer called allolactose binds to repressor so RNA pol. can bind and express the enzymes to allow for lactose digestion
Lac operon:
- What is constitutive expression?
- What happens when glucose levels are low in some bacteria?
- Mutation where lacZ and lacY are always expressed
- [CRP-cAMP] increases which induces positive regulation so lactose is digested for energy
Regulatatory transcription factors:
- What are their 2 binding sites?
- Enhancer on DNA and other to recruit general transcription factors (activators and promoters) which are assembled near TATA box in promoter
What do general transcription factors do? (2)
- Attract RNA pol.
- Bind to silencers and repress transcription
What enzyme removes introns and binds exons?
Spliceosomes
What is the purpose of postranslational modification?
Regulates structure and function - can change conformation to activate/inactivate a protein
What does methylation of cytosines on CpG islands cause?
Heavy methylation = no transcription
- this can be as a response to environmental signals
What does epigenetics change?
The way DNA is packaged which affects gene expression
What is an example of gene expression occurring on a chromosomal level?
One X chromosome out of the two in females is inactivated by becoming covered in Xist RNA from the X-chromosome inactivation centre
What happens to transcription when antisense RNA binds to some bacterial genes?
Prevents it
What are riboswitches?
Regulatatory sequences of mRNA that bind to ribosomes and change secondary structure which alters gene expression as translation cannot occur
2 players of human genome project?
- Public effort, international spread of chunks of genome and stored in yeast mini-chromosomes
- Celera genomics (private) - whole genome shotgun strategy
6 steps of public human genome sequencing strategy?
- DNA is partially digested and overlapping fragments are cloned in bacteria
- Large-insert clones analysed for markers or overlapping restriction sites
- Allows large-insert clones to be assembled into a contig (continuous DNA stretch)
- Subset of overlapping clones that cover entire chromosome are selected and fractured and cloned
- Small-insert clones sequenced and overlaps in these are used to assemble them in the right order
- Final sequence made by putting sequences of large clones together
How does whole genome shotgun sequencing work?
Small-insert clones are prepared directly from genomic DNA and are sequenced in an automated way
3 subsections of genomics?
Structural
Comparative
Functional
Structural genomics:
- What is the genome made up of?
Tandem repeats (highly conserved), transposable elements (can change position on genome), heterochromatin (condensed DNA) and non-conserved sections
Comparative genomics:
- Purpose?
- 3 sections of predicting function from sequence?
- Compares genomes to understand gene and species evolution, identify conserved and functional areas, finding how species relate and the function of genes
- Homologous, orthologs, paralogs
What is meant by homologous?
Evolutionary related genes
What are orthologs?
Homologous genes in different species from the same gene in a common ancestor
What are paralogs?
Homologous genes from duplication of a gene in the same organism
Comparative genomics:
- How can specific phenotypes be identified as having a role in evolution?
- What does it mean when genes show low rates of evolution?
- Genes that show accelerated evolution in evolution in lineages with specific phenotype
- Probably have a key function
Functional genomics:
- What does it determine?
- What is the transcriptome?
- What is a proteome?
- Gene function
- All RNA molecules transcribed from genome
- All proteins encoded by genome
Functional genomics:
- What is transcriptomics?
- 2 ways of doing it?
- Assessment of gene activity of many genes at a time
- Microarrays (nucleic acid hybridisation, known DNA fragment used as probe to find complimentary sequence)
- Next generation sequencing (RNA fragments sequenced then annotated by aligning them to a reference genome sequence)
What is cell division for?
Growth, cell replacement, healing and reproduction
How is bacterial genome organised?
Highly expressed genes near origin of replication and genes with similar functions are clustered together
Purpose of meiosis, meiosis I and meiosis II?
Meiosis = produces haploid gametes
Meiosis I = separation of homologous chromosome pairs
Meiosis II = separation of sister chromatids
3 causes of variation?
Crossing over, independent segregation and fertilisation
How do prokaryotic cells divide?
Binary fission (intitiated at origin of replication):
1. 2 plasimids form (each bound to different part of membrane)
2. Cell elongates and plasmids separate
3. Constriction at midpoint and division
4 parts of interphase in eukaryotes?
G1 phase = prep for DNA synthesis
S phase = synthesis of DNA
G2 phase = prep for mitosis, increase in size
G0 phase = nondividing
What does karyotype mean?
Number and shapes of chromosomes in a species
Phases of mitosis and meiosis are in week 5 reading
Prophase, prometaphase, metaphase, anaphase, telophase, prophase I, prometaphase I, metaphase I, anaphase I, telophase I etc
How does cytokinesis work?
Actin filaments form a contractile ring and the cytoplasm is pinched (animals)
In plants a phragmoplast is formed to form the cell wall
What do cyclin-CDK (cyclin dependent kinases) complexes control?
- Passage through the cell cycle (known as proteins seem to appear and disappear in cell cycle as they are activated and inactivated) — CDKs are only active when bound to cyclin
- They bind to a target protein and phosphorylate it to promote cell division and activate transcription factors in S phase
Role of cyclin B-CDK?
Prepares cell for mitosis
Role of cyclin D and E-CDK?
Prepare cell for DNA synthesis
Role of cyclin A-CDK?
Initiates DNA synthesis and prevents DNA replicating more than once in a cycle
3 checkpoints in cell cycle?
- Spindle assembly checkpoint (checks if chromosomes are attached to spindle)
- DNA damage checkpoint (checks if DNA is damaged)
- DNA replication checkpoint (checks if all DNA is replicated)
How is a tumour suppressed if DNA is damaged by radiation?
Protein kinase activated which phosphorylates p53 protein which acts as a tumour supressor by binding to DNA and turning on genes such as one to prevent G1 to S transition so DNA can be repaired (BUT p53 exposure over a long period increases Bax gene = apoptosis)
Why can cancer come from mutations in genes that control cell division?
- Loss of checks = uncontrollable division
- Proto-oncogenes (altered genes) have cell division role
Why is overactivation of one oncogene (possibility to cause cancer if activated) or inactivation of a single tumour suppressor not enough to cause cancer?
Requires greater build-up of mutations
How is division of cytoplasm different between sexes for gametes?
All products of meiosis in males become sperm, only one is an egg in females and the other 3 are polar bodies
- What is an oncogene?
- Protoncogene?
- A gene that causes cancer if it is mutated
- Mutated gene that causes cancer
3 ways mutations can be?
- Deleterious (leads to disease/disorders)
Neutral
Advantageous
What is mutagenesis?
Tools to understand genetic and developmental mechanisms
What are polymorphisms?
Changes in nucleotide sequence
How were mutations seen to arise?
Bacteria were plated, then half were picked up with a velvet disc and plated on another plate - one is a control and the other had a selective medium - some colonies lived meaning they were already mutated (did develop from exposure)
What are somatic mutations?
Mutations in non-reproductive cells that can cause cancer and are passed on in mitosis but aren’t heritable
What are germ-line mutations?
Less common mutations in gametes that can be passed onto half of next generation (but plants do not have a segregated germ-line)
Why do multiple mutations cause cancer?
Tumour suppressors are deactivated
When is the variation of mutation site and generation highest?
In small genomes
Why are most new mutations from males?
Sperm mutate much more than eggs as they divide much more
3 types of mutations?
- Substitution of a base (point mutation/SNP)
- Insertion of bases
- Deletion of bases
(insertion and deletion = indels or frameshift)
Point mutation:
- What causes it?
- 2 types?
- What does it not change?
- Why is location relevant?
- Wobble
- Transition (purine to purine or pyrimidine to pyrimidine) and transversion (purine to pyrimidine or pyrimidine to purine) in 2:1 ratio in rate
- Gene size
- If it occurs in introns there is no effect, if it is in exons there will be predictable consequences
Insertion and deletion:
- Why is indel difficult to use?
- What fraction of point mutation rate is the indel rate?
- Does not differentiate between insertions and deletions
- Between 1/10 and 1/20
What is the size of the region duplicated or deleted in copy-number variation?
One of more complete genes
Why tandem repeats/microsatellites used in forensics?
Change rapidly
5 causes of mutations in DNA?
- Single-stranded break
- Double-stranded break
- Missing bases
- Cross-linked T bases caused by UV radiation
- Bulky side group attached to a base
What is strand slippage?
New strand loops out and a nucleotide is added to new strand, then template strand loops out and one nucleotide is left out from new strand which results in insertions or deletions
What does unequal crossing over cause?
One product has insertion, the other has deletion
What is repeat expansion?
A special insertion whereby there is an increase in copies of groups of nucleotides
What is the mechanism of repeat expansion?
- DNA has many copies of a triplet
- Strands separate and replicate
- Hairpin forms on newly synthesised strand and part of the template will be replicated twice which increases the number of repeats on the new strand
- Two strands of new DNA separate and the strand with extra CAG copies is the template for replication
- Resulting DNA has 5 additional copies of CAG repeat
4 ways of repairing DNA damage?
- DNA ligase sealing breakage in sugar backbone (uses ATP to join 3’ OH and 5’ phosphate)
- Post-replication mismatch repair (single misplaced base is recognised by Muts proteins is removed and replaced by DNA segment using DNA polymerase, MutL, MutH and exonuclease)
- Base excision repair (incorrect base and sugar removed by AP endonuclease, and replaced)
- Nucleotide excision repair (multiple mismatched bases in a region recognised, enzymes to cleave DNA signalled, area of damage removed and replaced)
Difference between forward and reverse mutation?
Forward = wild (model) to mutant type
Reverse = mutant to wild type
What are:
- Silent/synonymous mutations?
- Missense/non-synonymous mutations?
- Nonsense mutations?
- Codon to synonymous codon
- Amino to different amino acid
- Sense to nonsense (STOP) codon
- How do different types of mutations affect fitness?
- Why is not that simple?
- Deleterious = decrease, can be lethal
Advantageous = increase
Neutral = no effect - Can be dependent on genetic background = epistasis
4 types of mutations by phenotypic effect?
- Loss-of-function mutations
- Gain-of-function mutations
- Conditional mutations (effect depends on another factor)
- Suppressor mutations (hides/suppresses previous mutation)
3 types of chromosome mutations?
- Duplications (most intra-chromosomal)
- Aneuploidy (1 chromosome has >2 copies)
- Polyploidy (all chromosomes have 2> copies)
- What is tandem duplication?
- Displaced duplication?
- Reverse duplication?
- Duplicated regions are adjacent to one another (special copy-number variation)
- Duplicated regions are separated by a non-duplicated region
- Segment flipped so genes are in opposite order to before
Why do duplications affect phenotype?
Unbalanced gene dosage
What can deletions cause (2)?
- Unequal crossing over
- Loop inpairing
What are inversions?
Segments of DNA breaking off and reattaching in the reverse orientation
Difference between paracentric and pericentric inversion?
Pericentric affects the centromere, paricentric does not
3 effects of inversions?
- Genes are split
- Position of genes affected which affects expression
- Meiosis is disrupted (loops change crossing over products so one has no centromere (this is lost) and the other has two - genes missing from gametes produced
What is translocation?
Material moves from one chromosome to another non-homologous chromosome (mostly reciprocal)
What is robertsonian translocation?
2 acrocentric chromosomes (1 long arm one side of centromere and 1 short arm the other) do translocation to produce 1 metacentric (normal) chromosome and a small-armed one that is not viable
- 4 types of aneuploidy?
- Example?
- Nullisomy (homologous pair lost, 2n-2)
- Monosomy (single chromsome lost, 2n-1)
- Trisomy (single chromosome gained, 2n+1)
- Tetrasomy (homologous pair gained, 2n+2)
- Down syndrome (91% nondisjunction in maternal meiosis, 7% paternal)
3 causes of aneuploidy?
- Deletion of centromere in mitosis/meiosis
- Robertsonian translocation
- Nondisjunction in mitosis/meiosis
What is the difference in what is made from nondisjunction in mitosis vs meiosis?
Mitosis = monosomic and trisomic cells
Meiosis = 2 gametes with extra chromsome,
2 gametes without
Why are older mothers more likely to produce children with down syndrome?
Higher aneuploidy due to reduced cohesin which means that chromosomes do not always go to right side so they are not divided fairly
Why is polyploidy mainly found it plants not animals?
Animals can only develop this if they reproduce parthenogenetically (fertilise themselves) which is unusual
Difference between autopolyploidy and allopolyploidy?
Autopolyploidy = all chromosomes from single species
Allopolyploidy = chromosomes from 2 species
How can autopolyploidy arise?
Nondisjunction in mitosis to form tetraploid cells if no cell division occurs, if it occurs in germline (meiosis) and diploid gametes form a triploid offspring is made
What affects phenotype?
Genotype and environment
What factors affect the effect of a mutation?
- If it is homozygous or heterozygous
- Genotype
- Environment