Bacterial Genetics

Question 1

What is genomics?

Answer 1

The study of genomes and DNA/RNA, genome replication, gene expression, genetic variation and distribution

Question 2

What are the two main components of the bacterial genome?

Answer 2

Chromosome and mobile genetic elements (like plasmids or prophages)

Question 3

What is a plasmid?

Answer 3

Autonomously replicating circular DNA

Question 4

What is a Prophage?

Answer 4

Viruses integrated into the chromosome

Question 5

What can you do from a whole genome sequence?

Answer 5

Predict cell function. You can identify patterns and homologous to known genes and motifs

Question 6

Where are the most differences between different strains of genomes found?

Answer 6

In the MGE

Question 7

What is a single nucleotide polymorphism?

Answer 7

Error in DNA replication

Question 8

What do many Mobile Genetic Elements encode?

Answer 8

Virulence, antimicrobial resistance or host specific genes

Question 9

What can acquisition of MGEs lead to?

Answer 9

New bacterial variants with enhanced virulence or resistance or host range

Question 10

What are the three major pathways for the MGEs to move between bacteria?

Answer 10

Bacterial transformation, transduction and conjugation

Question 11

How does bacterial transformation work?

Answer 11

Bacteria dies and releases the viral DNA into the environment. The recipient cell takes up that DNA up from the environment

Question 12

How does bacterial transduction work?

Answer 12

Bacteria is either infected with a virus, or the virus in the genome is popped out (usually under stressful conditions)

Bacteria then undergoes the lytic pathway and lysed releasing the virus

Virus then binds to a new bacteria and injects their genes into the recipient

Question 13

How does bacterial conjugation work?

Answer 13

Plasmids encode for the conjugation ability. Two bacteria come into contact with one another and make a pore between the two that allows the plasmid to move across to the other bacteria

Question 14

What is a lytic pathway in bacterial genomics?

Answer 14

When a virus starts to dictate what happens in the bacteria, so it makes lots of copies of itself

Question 15

Give some features of mobile genetic elements

Answer 15

• autonomously replicating circular DNA - not essential to the host bacteria - easy to manipulate

Question 16

What size are mobile genomic elements?

Answer 16

2 kb to >100 kb

Question 17

What is the restriction site on a plasmid?

Answer 17

Where you can add DNA

Question 18

What is generalised transduction?

Answer 18

Temperate bacteriophages accidentally packages host bacterial DNA or plasmids into long phage particles and delivers it to new bacteria

Question 19

Can all DNA transfer into any bacteria?

Answer 19

No

Question 20

What does the restriction modification enzyme do?

Answer 20

Binds to a specific sequence on the DNA and digests it

Question 21

How many subunits do restriction modification enzymes have?

Answer 21

3

Question 22

How do bacteria protect themselves from self digestion?

Answer 22

Makes a copy of the restriction modification enzyme without the enzyme bit, which methylates that bit of DNA, marking it as ‘self’

Question 23

What are some environmental triggers for bacteria?

Answer 23

Nutrients, oxygen, iron, temperature, bacterial pheromones, mammalian cells / hormones

Question 24

What are bacterial pheromones used for?

Answer 24

How the bacteria speak to each other

Question 25

Where are many bacterial virulence factors soley expressed?

Answer 25

In Vivo or in conditions mimicking those found in Vivo

Question 26

Where is ‘in vivo ‘?

Answer 26

In the host(the body)

Question 27

What can you do in transcriptional genomics?

Answer 27

Change the conditions depending on what you’re interested in

Question 28

What are the steps in transcriptional genomics?

Answer 28

• grow bacteria in conditions of interest - extract bacteria - extract bacterial mRNA - convert mRNA to DNA - sequence the DNA and quantitate the numbers of transcripts of each genes

Question 29

Why do we manipulate bacterial genomes?

Answer 29

To make tools for industrial production of proteins and for studying bacteria or gene function

Question 30

What is the first step in cloning?

Answer 30

Cut two different pieces of DNA using a restriction enzyme, leaving matching sticky ends on both . Mix the two together and use lipase to seal the join

Question 31

What are the most common cloning vectors?

Answer 31

Plasmids

Question 32

What is a cloning region?

Answer 32

Target site for multiple restriction enzymes

Question 33

Can any DNA be cloned?

Answer 33

Yes

Question 34

How do you check which clones have worked correctly?

Answer 34

Culture the cells in conditions where only the gene you’ve taken up could grow (add the antibiotic if you’ve added an antibiotic resistant gene)

Question 35

How do you construct a knockout (5)?

Answer 35

1- clone virulence gene into ‘suicide vector’ plasmid
2- clone antibiotic resistance marker into the gene to disrupt it
3- transform into a bacterial cell
4- Recombination via RecA protein (rare)
5- place onto agar with antibiotic and select for the rare isolate that has the resistance marker

Question 36

What is CRISPR?

Answer 36

Genetic manipulation of eukaryotes

Question 37

What does CRISPR stand for?

Answer 37

Clustered
Regularly
Interspaced
Short
Palindromic
Repeats

Question 38

What is a knockout?

Answer 38

When one of the organisms genes are made inoperative (knocked out)