Lecture 6 - horizontal gene transfer (pre-midterm)

Question 1

Core genome

Answer 1

Set of genes found in every strain of a given bacterial species. In E. coli, approximately 450 genes

Question 2

Pan genome

Answer 2

Set of all genes found in all strains of a given bacterial species. In E. coli, approximately 55 thousand genes.

Question 3

Genomic island

Answer 3

Block of genes found in one strain of bacteria but not another; sometimes can be transferred between bacteria thru horizontal gene transfer

Question 4

Base pair difference between E. coli O157:H7 and K12

Answer 4

500 base pairs are present in O157:H7 that aren’t in K12

Question 5

Sources of DNA for horizontal gene transfer?

Answer 5

Transposons, genomic islands, phages, plasmids

Question 6

Where did the gene sequence for shiga toxin come from?

Answer 6

Encoded by a lysogenic phage (integrates into the host genome)

Question 7

What is the source of most toxins found in bacteria?

Answer 7

Genes encoded by horizontally transferred genes

Question 8

What is a PAI?

Answer 8

Pathogenic associated island; genomic island that can encode one of multiple toxins

Question 9

Three main methods of horizontal gene transfer

Answer 9

1. Transformation
2. Transduction
3. Conjugation

Question 10

Transformation

Answer 10

Direct uptake of naked/free DNA into bacterial cells

Question 11

Transduction

Answer 11

Bacteriophage mediated transfer of DNA (usually DNA from one host cell to another)

Question 12

Conjugation

Answer 12

Transfer of DNA using mating pili

Question 13

Competent bacteria

Answer 13

Bacteria that are able to take up naked DNA from their environment

Question 14

How is competence achieved in bacteria?

Answer 14

• natural competency
• artificially achieve by chemical (use Ca + low temps to make membrane brittle) or physical (voltage drops to punch holes) means

Question 15

Observations of Griffith’s experiments with streptococcus pneumoniae

Answer 15

Rough (non-virulent) strain was able to take up virulence factors from heat-killed smooth (virulent) strain

Question 16

Competence in Gram negative bacteria

Answer 16

Machinery related to type IV pilus from N. gonorrhoeae
- DNA receptor recognizes certain DNA
- pilus pulls DNA to the cell
- dsDNA degraded to ssDNA
- ssDNA imported into cell

Question 17

Competence in Gram positive bacteria

Answer 17

Uses a pseudo-pilus that performs a similar action to Gram neg pilus. Grabs dsDNA, brings in ssDNA

Question 18

Requirements of bacterial conjugation

Answer 18

• mediated by conjugation pili assembled by donor cell
• requires cell-cell contact

Question 19

What is F factor?

Answer 19

Fertility factor. Low-copy plasmid that encodes proteins for the conjugation pilus.

Question 20

Where was F factor discovered?

Answer 20

F+ E. coli cells

Question 21

Replication origins in conjugative plasmids

Answer 21

1. oriV: “vegetative” used for replication during the cell cycle
2. oriT: “transfer” used during DNA transfer (conjugation)

Question 22

Method of replication from oriV in conjugative plasmids

Answer 22

Similar mechanism to theta replication. Bidirectional

Question 23

Method of replication from oriT in conjugative plasmids

Answer 23

Similar mechanism to rolling circle. Unidirectional, sends a single strand to recipient cell

Question 24

General mechanism of conjugation

Answer 24

1. Pilus forms between donor and recipient, contracts to bring cells together
2. Bridge forms between donor and recipient
3. Conjugative plasmid nicked at oriT and begins sending ssDNA through the bridge
4. Transferred strand re-circularizes and replicates

Question 25

Hfr strains

Answer 25

“High frequency recombination”
Strains of E. coli in which the F plasmid has integrated into the chromosome

Question 26

What is unique about Hfr cells?

Answer 26

• capable of transferring large segments of the chromosome into recipient cells via conjugation
• entire chromosome can be transferred in about 100 minutes

Question 27

Generalized recombination

Answer 27

Requires that the two recombining molecules have a considerable stretch of homologous DNA sequences

Question 28

Site-specific recombination

Answer 28

Requires a short specific sequence recognized by the integrase/recombinase enzyme

Question 29

Example of site-specific recombination

Answer 29

XerC resolution of catenated chromosomes

Question 30

Recombination with RecA

Answer 30

RecA finds the homology between two strands and mediates strand invasion

Question 31

Example of gene transfer into eukaryotes

Answer 31

Agrobacterium tumefaciens
- causes crown gall disease in plants
- contains a tumor inducing plasmid (Ti) that can be transferred via conjugation to plant cells

Question 32

Transduction vs generalized transduction

Answer 32

Transduction: transfer of genes between cells using bacteriophage particles
Generalized: transfer of any gene between bacteria

Question 33

Mechanism of generalized transduction

Answer 33

1. Phage DNA inserted into bacteria
2. Host chromosome degraded
3. New phages assembled with host DNA instead of phage DNA
4. Host DNA phage infects a new host
5. Bacterial DNA integrated into new host genome