CONJUGATION/ TRANSDUCTION/ TRANSFORMATION

Question 1

Why are bacteria useful genetic tools?

Answer 1

• cheap
• grow fast
• avoid ethical implications

Question 2

What did U- tube experiment prove about recombination?

Answer 2

need cell to cell contact

Question 3

Auxotrophic mutation

Answer 3

mutation that causes nutritional requirement not present in wild type

Question 4

Hfr strain

Answer 4

high frequency recombination strain with an F plasmid integrated into chromosomal DNA

Question 5

What is an F plasmid?

Answer 5

codes for genes that make pilli and carries antibiotic resistance genes

Question 6

What is an F’ plasmid?

Answer 6

If insertion sequences flank the F plasmid, they can recombine (but also carry some chromosomal DNA)

Question 7

Why do Hfr strains not always convert recipient cells to donor cells?

Answer 7

bc the plasmid is integrated into the chromosome, the connection will often be lost before there is to transfer the plasmid, the chromosomal DNA and then the last bit of plasmid

Question 8

How does conjugation occur?

Answer 8

• one strand of plasmid is nicked at the oriT
• F plasmid codes for the pilli formation
• 5’ end of free strand goes into the recipient cell and is synthesised discontinuously
• remaining strand is remodelled using rolling circle replication, therefore both cells retain/ gain a plasmid respectively

Question 9

What do interrupted mating experiments tell us?

Answer 9

circularity of bacterial chromosome and that the order of gene transfer are circular permutations

Question 10

How many crossovers are required to recombine a donor gene into a circular recipient chromosome?

Why ?

Answer 10

2

• one crossover produces an oversized liner genome so cell will die
• whereas 2 crossovers are requited to recombine a donor gene into a circular recipient chromosome

Question 11

In interrupted mating, why do those genes transferred early produce higher yields?

Answer 11

reduced risk of mating pairs breaking apart

Question 12

What factors influence mapping by recombination frequencies?

Answer 12

• distance between gene markers

- spontaneous separation of pilli between cells

Question 13

Which markers must we select for in mapping by recombination frequencies?

Why?

Answer 13

LATE

• ensures all the recombinants we look at have not been disrupted by spontaneous disruption of pilli between cells

Question 14

Once a late marker has been selected for, how do we produce recombination frequency maps?

Answer 14

determine how many early markers have been incorporated

Question 15

How did the U- tube experiment using Salmonella prove the phenomenon of transduction?

Answer 15

Bacteriophage particles can transfer through fine filters, which can then inject their DNA into bacteria

Question 16

Describe the process of generalised transduction

Answer 16

• phage particles inject their viral genome into a host
• phage replication occurs and occasionally recombination may occur which means that the phage may incorporate chromosomal DNA into viral genome in phage head preparation
• recipient receives chromosomal DNA, NOT viral genome so that the particles cannot lyse

Question 17

Transduction occurs rarely, when phages are at a ___ multiplicity of infection

Answer 17

LOW

Question 18

What is cotransduction?

What can it be used for?

Answer 18

When 2 markers close to eachother are both packaged in bacteriophage heads and injected into new bacteria

• can be used to find the order of markers

Question 19

What is a selection?

Answer 19

pick the one you want

Question 20

What is a screen?

Answer 20

Screening a particular trait?

Question 21

What is DNA transformation?

Answer 21

simply the uptake of DNA on surface of bacteria via binding

- recombination then occurs

Question 22

How can you make bacteria artificially competent for genetic transformation ?

Answer 22

• Treatment with ions or organic compounds

Question 23

Selecting for late markers ensures what?

Answer 23

all the recombinants we look at have not been disrupted by spontaneous disruption