Module 2--Inheritance & transfer of genetic material Flashcards
What are the properties of the genetic material?
-Genetic material must replicate, control the growth and development of the organism, and allow the organism to adapt to changes in the environment
What is the genotypic function of genome?
Replication
What is the phenotypic function of genome?
Gene Expression
What is the evolutionary function of genome?
Mutation
What is monoploid (x)?
One copy of the genome = number of unique chromosomes in a single complete set
What is haploid (n)?
Number of chromosomes in a gamete of an individual (sperm and ova)
What is diploid (2n)?
Two copies of the genome
What organisms are monoploid?
Bacteria and viruses
What is plasmid?
It is autonomously replicating circular DNA molecules that can replicate independently of the main chromosome in an extrachromosomal state
Why are bacteria and viruses good systems for studying nucleic acid movements and genome change?
Advantages of bacteria and viruses: -Small size -Rapid reproduction -Selective media -Simple structures and physiology -Genetic variability -Complete genome sequences
What is the life cycle of bacteria?
1) Growth phase 2) Duplication of genome 3) Division of cell and fission of cytoplasm
What are the phenotypes in bacteria that could be used fo selection or observation?
-Colony colour and morphology -Nutritional mutants -Antibiotic sensitivity and resistance
What is prototrophs?
Wild-type bacteria that can synthesise everything they need to grow and reproduce
How is genetic material transferred between organisms?
1) Vertically (parent to offspring) 2) Horizontally (between non-descendent/parent organisms of the same or different species
What is the direction of gene transfer in bacteria?
Unidirectional, from a donor cell to a recipient cell
What are the types of parasexual processes in bacteria?
-Transformation: uptake of free DNA -Conjugation: direct transfer of DNA from one bacterium to another -Transduction: a virus transfers bacterial genes from one bacterium to another
What is bacteriophage?
A virus that infects bacteria and archaea
What is the mechanism of transformation in competent bacteria?
1) A competent bacterium can bind exogenous DNA and transport it into cell 2) Exogenous DNA is bound to the receptor complex by the competence proteins ComEA and ComG. As the DNA is pulled through the channel composed of protein ComEC in the membrane by the ComFA DNA translocase, one strand of DNA is degraded by a deoxyribonuclease. This surviving DNA strand is stabilised by single-strand DNA-binding protein and RecA protein 3) The single strand of donor DNA is integrated into the chromosome of the recipient cell producing a DNA heteroduplex with different alleles in the two strands
What is the co-transformation?
It is the simultaneous transfer of two or more genes
What is the probability of co-transformation?
The close the two genes are on the host (donor) genome, the more frequently the two genes are co-transformed
What is the experiment that discovered conjugation?
Auxotrophic strains Y-10 (leu-, the-, thi-) and Y-24 (bio-, cys-, phe-) were grown individually (controls) and together in rich broth medium that contained all the amino acids and vitamins required for growth by both strains
What kind of bacteria are able to initiate transfer of conjugation?
Cells with fertility factor, which carry several genes required for conjugation to occur
What is TraG?
A membrane-anchored protein that forms a pore-like structure that could serve as a channel for translocation of the transferred DNA (T-DNA)
What is relaxase (Tral)?
It nicks the F factor at origin of transfer to initiate the process of DNA mobilisation and transfer
What is the mechanism of conjugation?
1) F pili of the F+ donor cell make contact with the F- recipient cell and pull the cells together. Genes on the F factor direct the synthesis of the conjugation bridge. One strand of DNA is cleaved at the origin of replication of the F factor by relaxase. 2) Rolling-circle replication transfers one strand of F factor into the recipient cell through the channel that forms between the conjugating cells. Replication of F factor occurs in both cells
What is the rolling circle replication?
The peeled-off strand is immediately regenerated using the remaining strand as a template
Why can’t F+ cells transfer more DNA to other F+ cells?
TraS, a protein, which is encoded by the traS gene on the resident F plasmid blocks TraG activity
What is an episome?
It is a genetic element that is not essential to the host
How can a bacterium obtain an episome?
It can either replicate autonomously or be integrated into the bacterial chromosome
What is high-frequency recombinant (Hfr) cell?
A cell with F factor integrated into the chromosome
What is insertion sequence or IS element?
It is transposable; can move around the chromosome; present in both episome and bacterial chromosome
How is F’ factor (F prime) formed?
F factor that integrated into the bacterial chromosome may also be excised by non-homologous recombination; this results in a portion of the chromosome being included in the excised F factor
What are the two types of bacteriophages?
Virulent (lytic) or temperate (lysogenic)
What are the characteristics of bacteriophage T4?
-Double-stranded DNA genome (single linear chromosome) -Protein head -Lytic/Virulent phage
What is the life cycle of the bacteriophage T4?
-T=0 min: T4 bacteriophage attaches to an E.coli cell and injects its DNA -T=2 min: Synthesis of phage-specific mRNAs begins -T=6 min: Replication of phage DNA begins; host DNA has been degraded by phage-encoded nucleases -T=14 min: DNA-filled heads, tails lacking tail fibres, and assembled tail fibres first appear -T=17 min: First intact phage particles are assembled -T=25 min: host bacterium is lysed, releasing progeny phage
How does T4 package its chromosome?
1) Replication of the injected T4 chromosome 2) Crossovers 3) Recombination within terminally redundant regions
What happens to the T4 genome when it it injected into a host cell?
It is terminally redundant and circularly permutated because of post-replication crossovers, concatamer cutting and headful packaging
What is headful mechanism?
Packaging DNA in a phage head, in which concatameric DNA is cut when the head is filled
What are the characteristics of bacteriophage lambda?
-Double-stranded DNA genome -Temperate (=may be lytic or lysogenic)
What is the life cycle of bacteriophage lambda in lytic pathway?
1) Injection of DNA into host cell 2) Replication of DNA 3) Synthesis of parts of phage 4) Assembly of phage 5) Host cell lysis
What is the life cycle of bacteriophage lambda in lysogenic pathway?
1) Injection of DNA into host cell 2) Site-specific recombination Once integrated, the lambda is called prophage
What determines the type of life cycle bacteriophage lambda is in?
Nutritional status of the cell
Where does site-specific recombination occur?
-Crossover occurs via homology between a 15 bp region of identity between att P and att B
How does site-specific recombination mechanism work?
1) Lambda integrase catalyses a recombination process that begins when several molecules of the integrase protein bind tightly to att P (on chromosome of lambda bacteriophage) 2) The resulting DNA-protein complex binds to att B (on bacterial chromosome), bringing the bacterial and bacteriophage chromosome close together 3) The integrase catalyses the required DNA cutting and resealing reactions, during a short region of sequence homology to form a tiny heteroduplex joint at the point of union
What is transduction?
A bacteriophage transfers DNA from a donor cell to a recipient cell
What is generalised transduction?
A random fragment of bacterial DNA is packaged in the phage head in place of the phage DNA
What is specialised transduction?
Recombination between the phage and the host chromosomes produces a phage chromosome containing a piece of bacterial DNA
Which phage can mediate generalised transduction?
Both lytic and lysogenic phage
Which phage can mediate specialised transduction?
Only lysogenic phage
What is the reversal of the site-specific integration process and its mechanism?
Normal excision of a lambda prophage at the att sites
What happens after the normal excision of a lambda prophage?
The phage enters lytic cycle and the cells produce more phage, then lyse and release the phage. The phage can then infect other bacteria
Where does anomalous excision of lambda prophage occur?
Occurs via Holliday junction migration
What is the mechanism of anomalous excision of lambda prophage and its product?
It produces specialized transducing phage carrying either gal or bio genes (= defective phage particles)
What is the mechanism of specialized transduction?
What are the criteria to distinguish between mechanisms of gene transfer?
- U-tube experiment
- Cell contact
- Sensitivity to DNase
What does the U-tube experiment prove?
It proves that DNA is protected inside the phage head (protein) and able to pass through the filter
What kinds of phages can become prophages?
Temperate phages, allowing their hosts to survive as lysogenic bacteria, and lysogenic phages (virulent phages cannot become prophages)
What are transposable elements–transposons?
DNA elements that can move around from one region of DNA to another
What are the different types of transposition?
1) Cut-and-paste transposition
2) Replicative transposition
3) Retrotransposition
What is cut-and-paste transposition?
An element is cut out of one site in a chromosome and pasted into a new site, using an enzyme called transposase
What is replicative transposition?
An element is replicated and one copy is inserted at a new site, one copy also remains at the original site
What is retrotransposition?
An element’s mRNA transcripts are used as a template to synthesize DNA molecules by reverse transcription, these are then inserted into new genomic sites
What kinds of transposons do bacteria have?
- Cut-and-paste transposon: insertion sequences (IS elements) and composite transpons
- Replicative transposons: Tn3 Elements
What kinds of transposons do Eukaryotes have?
Retrotransposons
What are insertion sequences (IS elements)?
Simplest bacterial transposons and their gene products are only involved in transposition
What is transposase?
A protein encoded by IS elements that is needed for transposition
What are terminal inverted repeats?
They are signals for recognition and cleavage
Insertion of an IS element causes:
Target site duplication (=a duplication of part of the DNA sequence at the site of the element)
What can target site duplication be used for?
It can be used as sequence scar to find IS elements in a genome
What is the function of IS elements?
Control expression of neighboring genes – polar or downstream affect
How do IS elements insert into chromosomes or plasmids?
1) Two strands of the target DNA are cleaved at different sites
2) IS element is inserted into the gap created by staggered cleavage of the target DNA
3) DNA synthesis fills in the gaps on each side of the IS element, producing a direct duplication of the target site
What are composite transposons?
They are bacterial cut-and-paste transposons (Tn)
How are composite transposons created?
They are created when two IS elements insert near each other
What are the charateristics of Tn3 elements?
- Larger than IS elements
- Contain genes that are not required for transposition
- Have simple inverted repeats at each end (not IS elements)
- Produce target site duplication when they transpose
What is the mechanism of the transposition of Tn3?
1) The transpsosase encoded by Tn3 catalyzes the formation of a cointegrate between the donor and recipient plasmids. During this process, Tn3 is replicated
2) Resolvase resolves the cointegrate by mediating recombination between the two Tn3 elements
What are the characteristics of conjugative R plasmids?
- Have spread multiple drug resistance in bacterial populations
- Can be transferred rapidly between cells in a bacterial population
- Have two components:
1. Resistance transfer factor (RTF): contains genes required for conjugative transfer between cells
2. R-determinant: contains genes for antibiotic resistance
What is the characteristics of Staphylococcus aureus?
-Quickly develops resistance to beta-lactam antibiotics
What is the mechanism of intrachromosomal recombination between transposons in the same orientation?
It produces a deletion
What is the mechanism of intrachromosomal recombination between transposons in opposite orientation?
It produces an inversion
What is the mechanism of unequal crossing over between transposons on sister chromatids?
It produces a gene duplication
