TRANSPOSONS Flashcards
What are transposons?
short pieces of DNA that can move around genome
What are transposases?
the enzymes that do so
Why are recognition sites so important?
repeats of 9 base pairs at each end that are used to transpose elements of DNA
What is IS10?
insertion sequences of around 22 base pairs at each end , 17 of which are identical in reversed orientation
How does Tn10 transpose?
- classic transposable element, has multiple copies of IS10 separate by a gene for tetraclycine resistance, used as recognition sequence for movement elsewhere
Tn10 is termed a ___ transposable element
composite
How can plasmids confer multiple drug resistance?
If drug resistance genes are found within insertion sequences.
What is a mutator phage transposable element like?
Can transpose sequence from one place to another randomly, which can cause mutations
Conservative transposition is also termed
- give an example
‘cut & paste’ transposition
- Tn10
How does ‘cut & paste’ transposition occur?
- 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
How does replicative transposition occur?
- 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
Important enzyme in Tn3 replicative transposition
Resolvase
Key point of replicative transposition
That the transposon not only stays where it is but moves as well
What happens in the intermediate of replicative transposition?
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
How do we identify the Shapiro intermediate?
inactivate TnpR resolvase gene
What does Mu not encode?
resolvase
Consequences of Mu not encoding resolvase?
loads of cointegrates produced, but no resolution phase as DNA integration
How do some do conservative and replicative simultaneously?
conservative as it keeps the same strands ‘cut and paste’ BUT there is replicative via replication and homologous recombination
Describe the Shapiro intermediate?
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.
