Bacterial Genetics Flashcards
Describe the structure of the bacterial genome.
All of the DNA in a bacterial cell in the bacterial genome.
It includes:
- chromosomes (single copy, circular, essential for life)
- mobile genetic elements (MGE) such as plasmids (autonomously replicating circular DNA) and prophages (viruses integrated into the chromosome)
How can we utilise a bacterial genome?
The genome determines what the bacteria is capable of. Bacteria can only do what their genome allows them to do.
From a whole genome sequence, we can predict cell function. We identify patterns and homology to known genes and motifs.
Putative Gene: It’s a segment of DNA whose protein, and its function, is not known, but based on its open reading frame, it is believed to be a gene.
Describe bacterial replication, and how it can lead to errors and evolution.
Replication of bacterial DNA is the first step in cell division. DNA polymerase enzymes catalyse the reaction to synthesise the new genome.
Sometimes, the DNA polymerase can make errors, such as single nucleotide polymorphisms (SNPs). Some errors will be fixed, while other not. They can accumulate.
Some errors can be advantageous, detrimental or neutral to the bacterial cell in a particular habitat. This is how bacterial cells evolve, through survival of the fittest.
We can use whole genome sequencing to reveal the evolutionary history of a species and how it is spreading.
What are mobile genetic elements (MGEs)?
They are genetic material that can move around within a genome.
What is the point of the horizontal transfer of MGEs?
Many MGEs encode virulence, antimicrobial resistance of host-significant genes. Aquisition of MGEs can lead to new bacterial variants with enhanced virulence, resistance or host range.
Describe the three ways in which MGEs are transferred across bacteria.
1) BACTERIAL TRANSFORMATION:
The donor cell has usually lysed, and had its DNA chopped up. This DNA is released and taken up by the recipient cell. The recipient cell may incorporate that DNA into its DNA, exchanging its genes for the other bacterium’s genes.
2) BACTERIAL TRANSDUCTION:
Here, bacteriophages (viruses of bacteria) are induced, they pop out and make phage particles. They then package phage DNA into these particles and release them (usually by killing the cell). The phage particles then inject their DNA into the recipient cell.
3) BACTERIAL CONJUGATION:
This is how big plasmids can move about. These plasmids encode genes needed to make a pore or pillus (basically, a tube) that connects the two bacteria (direct contact). The plasmid then replicates on its own and tranfers across.
Describe plasmids as MGEs.
Plasmids are a type of MGE. They are autonomously replicating circular DNA.
They are not essential for the host bacterium. Antimicrobial resistance genes in pathogenic bacteria are normally carried on plasmids.
Describe bacteriophages as MGEs.
Bacteriophages are viruses of bacteria.
They can either lyse bacteria or their genome just sits in the bacterial chromosome (prophage).
A prophage can encode important virulence genes.
What is generalised transduction?
It is when a temperate bacteriophage ‘accidentally’ packaged host bacterial DNA or plasmids into phage particles and delivers it to new bacteria.
Give two examples of bacterial immunity to protect themselves from foreign DNA (most importantly phage).
- Restriction Modification
- CRISPR gene editing
Describe Restriction-Modification (RM).
Instead of protecting themselves from the phage particles, they protect themselves from the phage DNA.
One mechanism that can be used is called Restriction-Modification (RM).
Restriction enzymes are made up of several units. In this example, the enzyme has a specificity subunit that binds the whole protein to a specific sequence of DNA. The restriciton subunit will restrict/cut through the DNA (both strands) at the specific site. There is a variant of the enzyme that doesn’t have the restriction subunit, only made up of the modification and specificity subunit. This variant binds to the same region of the DNA, and methylates the DNA sequence, and that protects it from the restrictive variant of the enzyme.
Describe CRISPR gene editing.
A phage injects its dsDNA into the bacteria.
An enzyme, CAS, will bind to the dsDNA and turn it into little bits of DNA which get integrated into the chromosome. These genes get integrated downstream of genes known as CRISPR.
The integrated genes can be transcribed into mRNA, and those mRNA fragments then get modified by other proteins intoprocessed cRNAs.
These cRNAs then combine with a third protein to form the CAS cRNA complex.
This complex can now bind to the dsDNA because of the specificity of the sequence. The third enzyme in the complex digests the DNA, deactivating it.
How is bacterial gene expression regulated?
Not all genes are expressed all of the time.
Also, we have regulators that can interfere with how RNA polymerase binds.
A classic example of gene regulation by a repressor protein is the lac operon.
Why do we manipulate genomes?
- to make tools for the industrial production of proteins
- to make tools for studying bacteria or gene function
Describe how we can clone genes using bacteria.
We take a plasmid and cut it at a specific sequence using a restriction enzyme. We then cut a piece of a human gene with the same restriction enzyme. This leaves bits of overhanging, linker DNA (sticky ends), which is used to bind the human DNA into a recombinant plasmid (via ligase).
The recombinant plasmid is then moved into a bacterium (most commonly used, E.Coli), which will then multiply and make lots of the protein encoded in the human gene.
