Chapter 5: Genes and genomes Flashcards
What is the genetic material of all organisms?
Deoxyribosenucleic acid / DNA
Define Gene
A heritible unit of DNA that encodes a product - either a protein or a functional RNA
What is a genome?
The entire genetic material of an organism
What are the key features of DNA that make it suitable to be heritable instructions?
- It has a stable structure
- Is able to accurately replicate (complementary base pairing and able to separate strands)
- Contains the information is a suitable form
Describe the structure of DNA
2 nucleic acid polymers, formed of nucleotides, wrapped around eachother in a right handed double helix, anti-parallel to eachother.
Each strand has a sugar-phosphate backbone (deoxyribose sugar and a phosphate group), and are connected through hydrogen bonds between each nucleotides nitrogenous base - either Adenine, Guanine, Thymine or Adenine.
What are the complementary base pairing rules in DNA?
Adenine (A) — Thymine (T)
Guanine (G) — Cytosine (C)
How many hydrogen bonds form between each complementary base pair found in DNA?
A - T: 2 hydrogen bonds
G - C: 3 hydrogen bonds
Thymine, Cytosine and Uracil are what type of bases?
Pyrimidines
Guanine and Adenine are what type of bases?
Purines
What is the structure of a purine?
A 6 membered nitrogen-containing ring fused to a 5 membered nitrogen-containing ring
What is the structure of a pyrimidine?
A six-membered nitrogen-containing ring
What is the name of the two strands of DNA in regards to DNA replication? What are the newly made strands called?
Template strand and Lagging strand (DNA)
Complementary strand (newly synthesized)
What is the function of helicase in DNA replication?
To separate the two DNA strands
What is the function of DNA polymerase in DNA replication?
To synthesise the next strand by catalysing polymerisation of nucleotides
What must be formed before DNA replication can take place?
Pre-replication complex
What is the approximate rate of error in DNA replication in humans?
1 in every million base pairs copied
Does DNA have an overall positive, negative, or neutral charge?
negative due to negative oxygen molecules on the attached phosphate group
What are the two types of grooves of DNA and what is their importance?
Major and minor grooves
- Allow access to the DNA for recognition and binding proteins
Draw a deoxyribose nucleotide!
Idk look it up :)
What type of bonding occurs between nucloetides and which areas bond?
- Covalent
- Between the sugar of one and the phosphate of the other
On which carbon of deoxyribose nucleotide is the phosphate group found?
5’
On which carbon of deoxyribose nucleotide is the -OH group found?
3’
What type of bonds form between the nitrogenous bases of DNA?
Hydrogen bonds
Which direction is DNA synthesised?
5’ –> 3’
The origin or replication contains high amounts of what type of base pairing? Why is this?
A-T pairings
Adenine and Thymine have 2 hydrogen bonds compared to the 3 between Guanine and Cytosine meaning the bonds will require less energy overall to separate the DNA strands.
What is the replication complex?
A protein complex containing a number of molecules and enzymes required for DNA replication. It binds to the origin of replication?
Where does the replication complex bind during DNA replication?
The origin of replication
What are replication forks?
The areas formed at either end of the replication bubble.
What is the function of helicase?
The separation of base bairs between the two DNA strands
Which direction does helicase travel down the DNA?
Bidirectionally
What is the function of single-stranded DNA binding proteins?
- Prevent base pairs from reannealing
- Protect DNA strands from nucleases that would break down the nucleic acid polymers.
What is formed when the two strands of DNA at the origin or replication are separated?
The replication bubble
What is the risk of supercoils forming?
Would eventually cause the prevention of strand separation and therefore prevent DNA replication.
What is the type of stress contained within a supercoil?
Torsional stress
What enzyme prevents supercoils that could prevent DNA replication?
DNA Topoisomerase
How does DNA Topoisomerase reduce the torsional stress of supercoils within DNA?
DNA Topoisomerase makes cuts in the sugar-phosphate backbone of a DNA-strand allowing the supercoils to unwind and therefore reduces the torsional stress. It then reseals the cuts.
What does Primase synthesise?
RNA primer
What is the function of RNA primer?
Faciliates the synthesis of DNA by RNA polymerase by providing an 3’ -OH group to begin synthesis from.
In which direction does Primase synthesise RNA primer?
5’ - 3’
Which specific DNA polymerase is involved in DNA replication
DNA Polymerase type III
Which direction does DNA polymerase read and synthesise?
Reads 3’ - 5’
Synthesises 5’ - 3’
What is the function of ligase in DNA replication?
Covalently bonds the DNA fragments/Ozaki fragments together
What are Ozaki fragments?
Short sections of the lagging strands newly replicated complementary strand.
‘DNA,RNA,DNA,RNA,DNA,RNA…’
What does pattern represent?
The lagging strands newly replicated complementary strands, showing the RNA primers and Ozaki fragments.
In DNA replication what is each strand of the parent DNA called?
3’ - 5’ is the leading strand
5’ - 3’ is the lagging strand
In what type of cells would replication bubbles eventually meet?
Prokaryotic
What can the polymerase chain reaction (PCR) be used for?
- DNA sequencing
- Gene cloning
What are the advantages of PCR?
- specific targeting of sequences
- Exponential inrease of yield
Describe the process of the polymerase chain reaction
- Denaturation | DNA is separated into two single strands
- Primer annealing | Oligonucleotide primers bind to their complementary sequences on the DNA strands
- Elongation | DNA polymerase initiates synthesis at primer, synthesises new DNA strands
What biomolecules are required to carry out the polymerase chain reaction?
- Double stranded template DNA
- DNA polymerase
- The four dNTPS (dATP, dGTP, dTTP, and dCTP)
- Two oligonucleotides (short single strands of DNA ~20 nucleotides long) that act as primers
What are the three steps of DNA replication?
- Initiation
- Elongation
- Termination
From what organism is DNA polymerase often extracted to use in PCR?
Thermus Aquaticus (Bacterium adapted to high temperatures)
What is mismatch repair (MMR)?
The repair system of DNA that can recognise and change incorrectly incorporated nucleotides in newly synthesises strands.
What carries out mismatch repair?
Mismatch repair enzymes
How do mismatch repair enzymes recognise the newly synthesises strand in some bacteria?
By its lack of methylation
Describe the process of mismatch repair?
- Complex of MMR enzymes cut new strand near distortion
- Unwinds short section
- Exonuclease degrades section of new strand starting at the cut, including the mismatch
- DNA polymerase synthesises new DNA in 5’ - 3’ direction using other strand as template
- DNA ligase joins newly synthesised section to rest of DNA
Done :)
What can the characteristic ‘molecular lesions’ of DNA be caused by? Give 2.
- Electromagnetic radiation | xyrays, gamma rays, UV light
- Toxic chemicals | Cigarette smoke, by-products of normal biochemical processes like reactive oxygen species.
What is deamination?
The removal of an amine group e.g., the deamination of cytosine to uracil
What is base excision repair (BER)?
Cellular mechanism of repair that removes a single damaged base from DNA and corrects it.
What kind of site would it be if a single base (not the sugar-phosphate backbone) was removed from a DNA strand?
Abasic site
Describe the process of Base excision repair (BER).
- ‘base name’ DNA glycosylase breaks bond between base and deoxyribose and removes damages base
- Sugar-phosphate unti is removed
- DNA polymerase adds new nucleotide
- DNA ligase seals single gap
Which bases can be affected by deamination?
Cytosine
Guanine
Adenine
What is depurination?
The complete loss of a guanine or adenine base leaving an abasic site.
Name two types of DNA damage that can be repaired by Base Excision Repair (BER)?
Deamination
Depurination
Name the three types of UV given out by the sun. Which is the most damaging to DNA?
UV-A, UV-B, and UV-C
UV-B is the most damaging to DNA
What can UV-B cause the formation of in DNA?
Pyrimidine dimers | Adjacent thymine or cytosine undergoing photochemical reactions leading to the covalent bonding of the bases, creating a kink in the DNA
What repair pathway is capable of reparing pyrimidine dimers caused by UV-B?
Nucleotide excision repair
Describe the process of nucleotide excision repair (NER)?
- Repair protein complex recognises kink in DNA
- Nuclease cuts the relevant DNA strand either side of incorrect base on the sugar-phosphate backbone
- DNA helicase unwinds the two DNA strand
- Cut section is released
- New DNA is synthesised for gap
- DNA ligase joins everything
What can cause double-stranded breaks (DSB) in DNA?
- Replication errors
- Highly energetic ionising radiation
If double-stranded breaks (DSBs) are not repaired, what will happen to the chromosome?
It will break/fragment
Describe the two mechanisms of repair for double-stranded breaks (DSBs) of DNA?
- Non-homologous end joining | Fragmented DNA has overreaching nucleotides at the location of the break removed. Ends are then ligated. This results in an overall loss of nucleotides.
- Homologous end joining | Damaged DNA pairs with the undamaged DNA from the homologous chromosome and the broken strand base pairs with the undamaged strands, new DNA is synthesised using template strands, ligase joints sections together, chromosomes separate.
Which repair mechanism for a double-stranded break (DSB) is most accurate?
Homologous end joining
Because homologous end joining uses a similar mechanism to genetic recombination in meiosis, what can be the result of this repair mechanism in terms of genetics?
Can result in cross-chromosomal genetic exchange :o
If many double stranded breaks exist, and many mechanisms of non-homologous end joining occur at the same time, what may happen?
Incorrect joining of DSB’s from:
- Different parts of the individual chromosome
-Between different chromosomes
Approximately how many base pairs exist in the human genome?
~ 3 billion
Approximately how many genes are in the human genome?
20 - 25 thousand
What is transcription?
The copying of information held in the base sequence of a DNA strand to an intermediate mRNA molecule.
Outline the steps of Transcription.
- Transcription factor protein complex binds to DNA at start of targeted gene.
- RNA polymerase binds close to complex
- Activator protein that is bound further along DNA, interacts with RNA polymerase
- RNA polymerase is released and travels down mRNA
- RNA polymerase unzips portion of double helix to expose bases
- One strand acts as template
- mRNA strand is synthesised using complementary base pairing
- mRNA molecule is released
What is Translation?
The process of using the information of DNA encoded into the intermediate mRNA to synthesise a polypeptide.
Where is the codon and anti-codon found? (translation)
Codons | mRNA
Anti-codons | tRNA
Where does translation take place?
Ribosomes
Outline the steps of translation.
- Codon read on mRNA
- tRNA with corresponding anti-codon binds to mRNA
- Peptide bond is formed between amino acids held by tRNA
- tRNA no longer needed released
- Repeat (elongation of polypeptide)
- Terminated at stop codon
- Folding of protein occurs through interactions between the amino acids
What is the genetic code?
The relationship in which a codon encodes for a single amino acid.
How many possible codons are there?
64 (4 x 4 x 4)
How many amino acids are there commonly found in human polypeptides?
20
How is the genetic code redundant?
Because some amino acids are encoded by multiple codons.
What is the start codon and what amino acid does it code for?
AUG | Methionine
What are the stop codons and what amino acid does it code for?
UGA, UAG, UAA | Does not encode for any amino acid
What is meant by the ‘Open Reading Frame’ (ORF)?
The section of DNA that encodes for a polypeptide
What could cause a single nucleotide to be changed in DNA?
- Deamination
- Depurination
- Misincorporation | Transitions or Transversions
- Deletions
- Insertions
What are the two types of misincorporations of nucleotides into DNA?
Transitions | Pyrimidine –> pyrimidine | or | purine –> purine
Transversions | pyrimidine –> purine | or | purine –> pyrimidine
What types of cells would have heritable mutations?
Germ-line cells (not somatic)
Is there higher mutation frequency in unicellular o multicellular organisms?
Unicellular
What types of mutations are there? (4)
- Missense mutation
- Nonsense mutation
- Silent mutation
- Frameshift mutation
What is a Missense mutation?
A change of nucleotide leading to a substitution of amino acid in the resulting polypeptide.
What is a Nonsense mutation?
A nucleotide change that results in a premature stop codon.
What is a Silent mutation?
A change in nucloetide that causes no change to protein sequence.
How can some mutations in DNA cause no mutations to the resulting polypeptide product?
The redundancy of the genetic code means that not all nucleotide changes cause a change in the amino acid encoded.
What is a Frameshift mutation?
When there is a shift in the reading frame that causes all amino acids after that point to be incorrect.
Why do most eukaryotic DNA mutations have no effect on phenotypes?
Because most DNA in eukaryotic organisms does not code for polypeptides, however some outside the protein coding sequences can affect gene function.
What do mutant recessive alleles generally represent in terms of function?
- Loss of reduction in function of gene, either because:
- reduced biochemical activity of encoded protein
- reduced protein expression
What type of gene mutations can cause loss of functions?
- Null mutations / amorphs | total loss of gene function
- Partial loss of function mutations / hypomorphs | in the name baby
Give 2 examples of DNA changes that may cause a loss of function?
- deletion of entire gene meaning no protein is synthesised
- nucleotide changes that remove the start codon, preventing translation
- nucleotide change than intoruduces stop codon too early
- Frameshift mutations causing incorrect amino acid sequence
- Nucleotide changes that may change amino acids integral to protein function
- changes in sequences involved in the control of gene expression may reduce the rate of protein synthesis
- transposable elements inserting themselves into a gene
What do mutant dominant alleles generally represent in terms of function?
A change or gain in function, either because:
-increased activity of gene product
-increased levels of transcription
-innapropriate [patterns of gene expression (when, where etc.)
- large chromosomal rearrangements that transport the genes coding region into proximinity with DNA transcriptional control sequences that normally activate another gene
What does a neomorphic mutation lead to?
A completely new function of the gene.
What are the predominantly studies prokaryotes relating to understanding of genomes?
E. coli
Bacteriophage
Describe the prokaryotic gene structure.
On the DNA template strand:
- Regulatory region
- Operon | group of genes with related function
- Transcription terminator
What is the function of the regulartory region in a prokaryotic gene?
- Regulates gene expression
- Contains binding sites for RNA polymerase and regulatory proteins which in turn interact with RNA pol to change efficiency of transcription
What type of mRNA would be transcribed from a prokaryotic gene operon?
A single polycistronic transcrip
What is an example of a prokaryotic operon?
lac operon | E. coli | included lacY, lacZ, and lacA - all proteins encoded are involved in the metabolism of lactose.
What is the structure of most prokaryotic chromosomes?
A single circular chromosome.
If a prokaryotic chromosome begins to gain torsional stress and supercoils, what is the solution?
Topoisomerase :D
What are plasmids?
Small circular extrachromosomal genetic elements in prokaryotes that replicate independently, have a single origin of replication, a small number of genes (which encode proteins to faciliate their own movement)
What are the implications of prokaryotic plasmids on human health?
- genes encoding toxins
- antibiotic resistance / antibiotic inactivating enzymes
- Infectivity
- virulence factors
How are plasmids transferred between bacteria?
Horizontal gene transer, specifically conjugation
What are the two different DNA replication mechanisms seen in bacterial plasmids, and which type of bacteria uses which?
Gram-negative bacteria | replicate bidirectionally
Gram-positive bacteria | rolling circle replication
Desribe rolling circle replication?
DNA replication of gram-positive bacterial plasmicds in which replication regins at the origin where one strand is nicked. The circle rolls diplacing the original strand and many plasmid lengths are synthesised, separated, and then undergo circularisation, ligation, and second strand synthesis is required.
What does the mitochondrial genome usually encode?
- mitochondrial tRNAs
- rRNAs
- some protein components of the electron transport chain
Where are the main differences seen between the ‘universal’ genetic code and the mammalian mitochondrial genetic code?
In the stop and start codons.
Whose genome is bigger, prokaryotes or mitochondria? Why?
Prokaryotes
Many of the prior mitochondrial genes are believed to have been transferred to the nuclear genome
How do prokaryotes pass their genes onto daughter cells?
Asexual binary fission
What are the three mechanisms of horizontal gene transfer between prokaryotic cells?
- Conjugation
- Transformation
- Transduction
What contributes to genetic variation in prokaryotic populations?
Horizontal gene transfer
What is an F plasmid?
A plasmid containing:
- an origin of replication
- oriT | origin of initiation of DNA transfer
- set of tra genes required to form sex pilus
How are E.coli strands containing, and ones not containing, an F plasmid known?
Present F plasmid: F+ (superscript +)
No F plasmid: F- (superscript -)
What are strains known as if the F plasmid has become incorporated into the chromosome?
Hfr (high frequency of recombination) strains
What does the amount of chromosomal DNA transferred during conjugation depend on?
The time that the transfer can be maintained
- breakage of pili happens very frequently as they are fragile
What is Transformation in terms of prokaryotic genetics?
The process carried out by some prokaryotes in which naked DNA is picked up from their environment and incorporated into it’s own genome.
How could you force an E.coli bacteria, which does not naturally carry out Transformation, to begin picking up and incorporating naked DNA into it’s genome?
Treat it with metal ions, increasing membrane permeability, becomes competent to take up DNA.
Other examples:
- electroporation
What is Transduction in terms of prokaryotic genetics?
The transfer of DNA from one cell to another carried out by a virus (bacteriophage).
What does ‘competency’ mean in terms of transformation in bacteria?
Competency is a measure of the permeability of the cell wall and cell membrane, and therefore the cells ability to take in extracellular DNA.
What are the two types of transduction?
Generalised | phage capsid accidently assembles itself around bacterial DNA, transferring it to a new hose
Specialised | Icorrect excision of DNA during induction means bacterial DNA fragments can be picked up and replicated, then transferred to new hosts
What is The C-Value Paradox?
A haploid genomes size does not correspond strongly to the organisms complexity.
What is the difference between the continuity of protein-coding regions in prokaryotes and eukaryotes?
prokaryotes have continuous protein coding regions
Eukaryotes have interrupted protein coding regions consisting of:
- Exons | coding sections
- Introns | Non-coding sections
When and how are introns removed from mRNA?
During post-transcriptional modifications
Splicing
Desribe the structure and parts of a eukaryotic gene.
- Distal regulatory regions
- Proximal promoter regions
- TATA box
- Exons
- Introns
-Transcription termination sequences
In a eukaryotic gene, what is the function of distal regulatory regions?
Binding site for regulatory factors that act upon signals from a cells environment
In a eukaryotic gene, what is the function of the proximal promoter region?
Binding site for protein factors that regulate transcription and promote the binding of RNA polymerase
What is the TATA box?
A region in a eukaryotic gene that acts as a binding site for RNA polymerase.
How can several forms of a protein be encoded by a single eukaryotic gene?
The presence of introns and exons, and selective splicing, means different combinations of exons can form the mature mRNA, therefore creating different forms of a protein.
Where are regulatory regions found in relation to a eukaryotic gene gene?
Upstream or downstream, close or far
Why might some genes have multiple copies per genome? Give an example of a gene where this may occur.
Frequent use
Need large quantities of encoded products
e.g., genes that encode histone proteins, rRNA
What are gene families?
Genes with similar sequences and have related, but distict, roles.
In terms of eukaryotic genome composition, what proportion is formed of exons?
1.5%
What are transposons / transposable elements?
Sections of DNA that can move around the genome
What is the function of transposons?
No specific function has ever been found.
Everything they do is for their own maintanace and for faciliating their transposition.
What are the two classes that use different mechanisms of transposition?
- Uses RNA as an intermediate which is then reverse transcribed by reverse transcriptase.
- Transposase cuts the transposon and inserts it elsewhere in the DNA
What is the most likely mutation to be caused by a transposon inserting itself into the ORF of a gene?
Loss of function.
What is Satellite DNA / Simple sequence repeats?
Short sequences that repeat, found clustered around structural features of chromosomes (i.e. centromete).
How can simple sequence repeats of DNA be separated from non-repetitive DNA?
Centrifugation | DNA of different denseties.
What is the problem with the lagging strand newly synthesised complementary strand at the end of DNA replication?
Cannot be copied at the extreme 3’ end due to there being no free 3’ -OH group once the RNA primer has degraded which leads to a slowly decreasing length of chromosome after each replication.
What exist in eukaryotic cells to overcome the shortening of the chromsome caused by problems with replicating the lagging strand during DNA replication?
Telomeres
What is a telomere?
A protective region found at the end of chromosomes containg thousands of repeats of a single sequence.
(TTAGGG)n in humans.
What enzyme is responsible for synthesising telomeres?
Telomerase
What type of cells would you expect Telomerase to be very active in?
Stem cells, germ-line cells, cancer cells
What happens if telomerase is not present in a cell?
It will undergo a set number of divisions before entering a cellular senescence state in which it is alive but can no longer proliferate.
What type of cells would you expect to have no telomerase activity?
Somatic cells
What do most viral genes encode?
Coat proteins to form the capsid that protects the genome
What are homolgous genes?
Genes across species that share some sequence similarity due to a common evolutionary origin.
What does much of the genetic variation in H. Sapiens take the form of?
Single nucleotide polymorphisms (SNPs)
- single nucleotide differences
-most cause no gene activity differences
Copy number variation (CNV)
- Variation in gene copies due to duplication of genes or larger chromosomal areas
- results in variation of gene expression
-may result in the evolution of gene families which are structurally related genes with similar or different functions
What is a gene family?
Genes share a similar nucleotide sequence that have diverged from an original ancestral sequence.
What mechanisms exist that can result in segments of DNA to be duplicated?
- DNA replication errors
- Unequal crossing over between a homologous chromosome pair during meiosis
- Transposons may accidently carry some neighbouring DNA with them
- Duplication of entire chromosome / entire genome
What types of chromosome rearrangement are there?
- Duplications | A section has been duplicated
- Deletions | A section has been lost
- Inversions | Section has been flipped in orientation different to normal chromosome order
What is segmental duplication? How common is it?
- Duplication of genome sections containing many genes
- Common
What is recombination frequency between two loci related to?
The distance between them
What can SNP’s linked to disease-related genes be used as?
Genetic markers to identify the presence of disease-related alleles
