TRANSPOSONS

Question 1

What are transposons?

Answer 1

short pieces of DNA that can move around genome

Question 2

What are transposases?

Answer 2

the enzymes that do so

Question 3

Why are recognition sites so important?

Answer 3

repeats of 9 base pairs at each end that are used to transpose elements of DNA

Question 4

What is IS10?

Answer 4

insertion sequences of around 22 base pairs at each end , 17 of which are identical in reversed orientation

Question 5

How does Tn10 transpose?

Answer 5

• classic transposable element, has multiple copies of IS10 separate by a gene for tetraclycine resistance, used as recognition sequence for movement elsewhere

Question 6

Tn10 is termed a ___ transposable element

Answer 6

composite

Question 7

How can plasmids confer multiple drug resistance?

Answer 7

If drug resistance genes are found within insertion sequences.

Question 8

What is a mutator phage transposable element like?

Answer 8

Can transpose sequence from one place to another randomly, which can cause mutations

Question 9

Conservative transposition is also termed

• give an example

Answer 9

‘cut & paste’ transposition

• Tn10

Question 10

How does ‘cut & paste’ transposition occur?

Answer 10

• transposase makes breaks on 2 opposite ends of DNA
• 3’ end of transposon is joined to 5’ end of cleaved target
• DNA synthesis fills in target and fills in gaps

Question 11

How does replicative transposition occur?

Answer 11

• Begin with 2 substrates, plasmid A with the transposon and plasmid B without
• cointegrate formed where replication has occurred, 1 copy of transposon points up and one downwards, in direct repeat with respect to eachother
• a site specific recombination by RESOLVASE occurs which

Question 12

Important enzyme in Tn3 replicative transposition

Answer 12

Resolvase

Question 13

Key point of replicative transposition

Answer 13

That the transposon not only stays where it is but moves as well

Question 14

What happens in the intermediate of replicative transposition?

Answer 14

formation of a Shapiro intermediate

• creation of 2 replication forks as the 3’ end of transposon is linked to the 5’ end of new piece of DNA
• BUT the 5’ end of transposon remains attached to the ORIGINAL DNA
• allows retention in the original plasmid

Question 15

How do we identify the Shapiro intermediate?

Answer 15

inactivate TnpR resolvase gene

Question 16

What does Mu not encode?

Answer 16

resolvase

Question 17

Consequences of Mu not encoding resolvase?

Answer 17

loads of cointegrates produced, but no resolution phase as DNA integration

Question 18

How do some do conservative and replicative simultaneously?

Answer 18

conservative as it keeps the same strands ‘cut and paste’ BUT there is replicative via replication and homologous recombination

Question 19

Describe the Shapiro intermediate?

Answer 19

this is formed during replicative transposition. The 5’ end of the insert remains attached to its original plasmid DNA sequence whereas the 3’ associates with the new piece., creating 2 replication forks.