Lecture 3: Bacterial Genetics and Gene Transfer Flashcards
Are prokaryotes haploid or diploid? Why?
Haploid. Because they (generally) have 1 circular chromosome (0.5-14Mbps)
What are the two forms of genetic material in prokaryotes
Plasmids (extrachromosomal DNA) and chromosome
What are plasmids?
Usually non-essential, small and self replicating, confer a level of adaptive advantage
What is the nucleoid?
Region of cytoplasm containing genetic material
How are genes packaged in prokaryotic genetic material?
More condensed and close together. Grouped into units called OPERONS - which are under control of a single regulatory sequence
What is an operon and how is it transcribed
An operon is a collection of genes all controlled by the same regulatory sequence. An operon is transcribed into a single mRNA molecule encoding several proteins - “POLYCISTRONIC”.
What does polycistronic mean?
A single (mRNA) molecules encoding several proteins
does prokaryotic mRNA undergo posttranscriptional modification or have introns?
No and no
How is it that transcription and translation are able to act almost simultaneously (coupled) in prokaryotes?
Due to the lack of nuclear barrier alongside no posttranslational modifications
Prokaryotes VS Eukaryotes
Polycistronic monocistronic
no introns introns
no post-trans. mods mods req. for translat.
coupled uncoupled
Why is the prokaryotic genome highly adaptable and quick evolving? Explain what it is.
horizontal gene transfer (HGT). The movement of genetic material between two related/unrelated cells. (combined with spontaneous mutations)
Opposite of vertical gene transfer
Outside of direct cell to cell transfer, how else can HGT occur?
Transformation
Conjugation
Transduction
Give examples of adaptations that are the result of HGT, contributing to bacteria’s rapid ability to evolve
- ability to use new substrates as food
- antibiotic resistance
- detoxifying harmful chemicals (heavy metals, salts…)
- virulence
What is transformation?
Direct uptake of extracellular ‘naked’ DNA by bacterial cells.
DNA usually comes from nearby degraded cells. Can be in the form of chromosomal fragments or plasmids
This DNA can then be degraded for materials for own metabolism or maintain by ingestion in the chromosome or independent replication (plasmids).
What is “competence”? What are the two types?
The physiological state allowing acquisition of exogenous DNA.
- Natural
- Induced
What is Natural competence?
Genetically encoded capability to uptake and incorporate exogenous DNA.
Possess genes encoding machinery for capture, transport, and incorporate into their chromosome.
How many species are natural competent? Gram + or - bacteria?
> 80 species. Including both gram + and - bacteria
When and by who was natural competence first seen and with what bacteria?
1928, Frederick Griffith, Streptococcus pneumoniae strains
Briefly explain Frederick Griffiths experiment
S. pneumoniae has two strains; rough (R) which is non-virulent and smooth (S) which is virulent. Both was injected into mice. R = no death, S = death of mice. Next…
Injecting mice with flame killed S lead to NO death, however, injecting mice with both healthy R alongside flame killed S did lead to mice death as the R was transformed to virulence.
What is induced competence? How is it done?
Artificial permeabilization of bacterial cell envelope.
- Heat shock (treating with chemicals like CaCl2 before hand)
- Electroporation
What is conjugation?
Mechanism of DNA transfer involving direct cell to cell contract via formation of specific structures called CONJUGAL PILI
Main transfer mechanism for plasmids
By who, when, and how was conjugation described?
Joshua Lederberg and Edward Tatum in 1946. Noticed E.coli mutant cells with impaired ability to produce some nutrients (autotrophs) were able to produce them after being in contact with other E.coli cells possessing the ability to do so (phototrophs).
What is the process/mechanism of conjugation?
- DONOR (F+) cell produces a pilus attaching to RECEPTORS in RECIPIENT (F-) cell
- Pilus then retracts bring the cells closer, establishing a RELAXOME BRIDGE
- Plasmid is nicked at specific location called “ORIGIN OF TRANSFERENCE” (oriT). One of its strands transfers through the relaxosome.
- After reconsititution of the plasmid, RECIPIENT CELL BECOMES A POTENTIAL DONOR
What is an “episome”
When a plasmid integrates into a chromosome
Can many plasmids be present in one cell? when cant they?
Yes. When they possess the same replication machinery - classified as the same INCOMPATIBILITY GROUP
Give an example of where conjugation is present
Plant pathogen Agrobacterium tumefaciens. It is a tumour inducing plasmid, also containing the genes required for conjugation called T-DNA. When it the bacterium penetrates the plant through a wound, T-DNA is transferred into the plant cell nucleus. Result in expression of hormones stimulating cell growth and tumour formation
What has the Agrobacterium allowed
the ability to genetically manipulate plants
What is transduction
Transfer of DNA from one bacterial cell to another mediated by bacterial viruses, bacteriophages. DOESNT REQUIRE DIRECT CELL-CELL CONTACT.
When the DNA is impanted, 1 of 3 things can happen:
- genetic material identified and destroyed
- phage genome hyjacks machinery and starts multiplying - LYTIC CYCLE
- phage genome integrates in chromosome gorming a PROPHAGE, remains dormant or at low level of expression - LYSOGENIC CYCLE
What are the 3 main types of transduction
- Generalised transduction - transfer of DNA from any part of host genome to recipient cell
- Specialized transduction - transfer of a few specific sets of genes between cells
- Later transduction - seen in S. aureus, sim to specialized but can transfer more host DNA at higher frequency rates
Where can you find more detail on general, specialized and lateral transduction?
Slides
What are the applications of transduction?
- Capsids as delivery vehicles
- Making DNA libraries
- Transfer DNA between cells
- Engineering integration machinery into plasmids, allows stable integration
