Flashcards: Bacterial Genetics (Pathology)

1)A unit of heredity located on a specific region of the DNA molecule

2)Genes are involved in the production of a particular product through its expression

- Putative genes encoding proteins with predicted functions (eg. toxins, virulence factors, metabolic pathways)

- Gene regions with predicted functional regions (eg. Transmembrane region, ATP binding region)

- Start and stop regions for those genes

- Regions where gene regulators and RNA polymerase bind

- Integrated mobile genetic elements

Putative

Variations at a single position in a DNA sequence among individuals within a population, involves the substitution of a single nucleotide (A, T, C, or G) for another in the DNA sequence

Whole Genome Sequencing

The evolutionary history of a species and how it is spreading

The UK Midlands in 1986

In various organisms, including bacteria, archaea, and eukaryotes

Yes

2 kb - 100 kb

No

Via plasmids

Can lyse and kill bacteria OR have its genome sit in the bacterial chromosome (prophage)

It is a measure of the severity of the disease that the microorganism can cause in an infected host (pathogenicity)

No

The lac operon

In vivo

Genes important for survival and virulence are expressed

Isolate the Gene of Interest:

Extract and purify the DNA containing the gene of interest

Select a Plasmid Vector:

Choose a suitable plasmid vector based on size, replication properties, and any specific features required for downstream applications

Digest the Plasmid and Gene DNA:

Use restriction enzymes to cut both the plasmid vector and the gene of interest. This creates compatible ends for ligation

Ligate Gene into Plasmid Vector:

Use DNA ligase to join the cut ends of the plasmid and gene, creating a recombinant DNA molecule.

Transform Bacterial Cells:

Introduce recombinant plasmid into bacterial cells through transformation

Select for Transformed Cells:

Culture the transformed bacteria on agar plates containing an antibiotic that the plasmid carries a resistance gene for. This ensures only cells with the plasmid survive

Identify Positive Clones:

Screen for bacterial colonies that contain the gene of interest in using methods like PCR or restriction enzyme digestion

Amplify and Extract Plasmid DNA:

Culture and amplify the positive clones, then extract and purify the plasmid DNA

Insert the virulence gene into a "suicide vector"

Integrate an antibiotic resistance marker into the target gene to disrupt its function

Introduce the modified plasmid into a bacterial cell through a process called transformation

Encourage recombination using the RecA protein, although this process is not very common

Plate the transformed cells onto agar containing the antibiotic, selecting for the rare isolates that successfully incorporated the resistance marker

-Is a technique used to study the function of a specific gene by rendering it nonfunctional

-Researchers can observe the effects on an organism and understand the role that gene plays in various biological processes

A plasmid lacking the origin of replication and therefore cannot replicate independently

To introduce foreign genetic material into a cell, allowing it to take in and express the inserted genes

Clustered Regularly Interspaced Short Palindromic Repeats

40%

Provides Adaptive immunity or protection from foreign DNA or MGE that have been previously encountered

New technologies change everything. e.g. sequencing, bioinformatics, gene expression technologies; discovery of the toolbox of restriction enzymes, plasmids, CRISPR, etc.

We have learned -

How genomes are – replicated, vary, transferred, expressed, modified...How bacteria colonise, spread, cause disease, can be diagnosed, treated or prevented...How to make artificial proteins, structures, cells and in industrial quantities.How to genetically manipulate cells for study, therapeutics and industrial applications.