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Hilary Term 1 > Prokaryotic Gene and Sex Exchange > Flashcards

Prokaryotic Gene and Sex Exchange Flashcards

1
Q

Bacteria have 3 main ways of acquiring DNA

A

transformation, transduction and conjugation

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2
Q

Describe transformation

A
  • common amongst gram positive bacteria
  • acquisition of DNA directly from the environment
  • very powerful for genetic exchange
  • acquiring new functions such as antibiotic resistance
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3
Q

During transformation, DNA acquired must be

A

recombined into the host, except if it is a self-replicating plasmid

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4
Q

Transformation occurs by

A
  • nicking of one DNA strand
  • a single stranded binding protein attaches
  • allows cross strand exchange between the DNA chi
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5
Q

What nicks the DNA strand during transformation?

A

an endonuclease

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6
Q

What is a splice in transformation?

A

When the nick is on the external strand of the chi

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7
Q

What is a patch in transformation?

A

when the cut is on the internal strand of the chi

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8
Q

Transformation can be monitored in the lab using

A

a marker that tracks the path of recombination

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9
Q

Trp- means a bacteria

A

can’t grow on Trp

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10
Q

Plating cells with no Trp and seeing when they begin to grow proves that

A

they have taken up some DNA

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11
Q

What does measuring DNA uptake require

A

an auxotrophic marker, or antibiotic resistance system

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12
Q

Described induction of transformation in gram negative bacteria

A
  • bacterial cells bind the DNA at the surface and strip it with nuclease
  • it enters the cell single stranded
  • recombination allows the cell to replicate
  • makes the cell very leaky
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13
Q

Inducing transformation in gram negative bacteria involves

A
  • electroporation
  • surface shielding of the extracellular polysachharide and S protein layers stops external molecules such as toxins, and antibiotics from invading,
  • stops immune attack.
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14
Q

Describe electroporation

A

an electric pulse knocks holes in the membrane

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15
Q

Give an example of a naturally transformable clade of bacteria

A

the Streptococci

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16
Q

Generalised transduction involves

A

aquiring DNA from a phage

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17
Q

Describe generalised transduction

A
  • a lytic phage enters the cell and replicates
  • some phage DNA combines with any part of the chromosomal or plasmid DNA
  • creates a transducing particle
  • transfers genes from one strain to another, from host to recipient cells
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18
Q

Describe the basics of specialised transduction

A
  • requires a transducing phage
  • activated by processes such as the LexA SOS cycle
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19
Q

Give an example of a transducing phage

A

lambda phage

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20
Q

Describe transducing phages

A

composed of a phage head and a linear molecule

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21
Q

Describe the specifics of specialised transduction

A
  • the transducing phage cyclises in the host, forming a sphericule with adhesive ends
  • site-specific endonucleases cut at ATBs
  • allows DNA insertion and integration between the galactose and biotin genes, forming a prophage
  • prophage can then excise and replicate if the lytic cycle is induced
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22
Q

ATBs

A
  • attachment sites in bacterial chromosomes
  • complementary to ATPs in the phage
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23
Q

Describe rolling circle replication

A
  • a way of making multiple copies of a genome
  • used by the lambda phage genome
  • catalysis occurs from 5’ to 3’
24
Q

Describe rolling circle replication in lambda phage genomes

A

can make hundreds of lambda phage genomes as it continuously replicates

25
Q

What is a ‘normal event’?

A
  • induction by the lytic phage
  • excision
  • rolling circle replication
  • transcription and translation of the capsid protein
  • cell lysis
  • phage release
26
Q

What is the function of the capsid protein?

A

encapsulates the DNA

27
Q

Describe a ‘rare’ event

A
  • the excising DNA takes another gene because the excision cut in the wrong place
  • creates a hybrid phage containing chromosome
  • phage loses some of its own DNA
  • only the genes very close to the phage are at risk of becoming hybridised
28
Q

Give an example of phage DNA in loss in a rare event

A

excision at the junction between ATB and ATP.

29
Q

Hybrid phages are

A

often defective

30
Q

‘Rare’ phages will therefore have been

A

mixed in with lots of wild phages

31
Q

What do wild phages provide?

A

the necessary proteins in transduction

32
Q

How are rare events isolated in the lab?

A

removal of the wildtype

33
Q

Describe the major constraint acting on transduction

A
  • capsid proteins have a 45-50kb packaging limit
  • if the package is either too small or too large, the phage head won’t fit
34
Q

Describe conjugation in bacteria

A
  • what causes the ‘sex change’
  • transfer of the F plasmid across the conjugation bridge
35
Q

Describe the F plasmid

A
  • contains the genes for the sex pillus
  • often recombines into the chromosome through insertion sequences
  • can exist as both plasmid and chromosomal DNA
36
Q

What is the OriT?

A

the transfer origin of the plasmid

37
Q

Give an example of an insertion sequence for the F plasmid

A

IS3 (insertion sequence 3)

38
Q

Integration direction of the F plasmid depends upon

A

orientation of the insertion sequence

39
Q

F episomes

A

transfer chromosomal DNA

40
Q

Describe wholesale transfer of E. coli

A
  • almost all of its chromosome can be transferred from one strain to another in 100 minutes
  • between Hfr strains
  • can be used to show the order of genes in the chromosome by interrupting the experiment at various intervals
41
Q

Describe classical genetic mapping experiments

A
  • landmark
  • use wholesale transfer
42
Q

Describe archaeal gene transfer

A

Archaea display a TraG-like homologue for stability
- mobile elements have inverted repeats at either end, and composite transposons with 2 sets of insertion sequences at eirther end (L and R)

43
Q

Name an archaeal mobile element

A

insertion sequence 2

44
Q

Name an example of the composite transposons contained within mobile elements

A

IS50R and IS50L

45
Q

IS

A

insertion sequence

46
Q

Conservative transposition involves

A
  • the transposon being excised from the donor and transferring to a new location
  • the donor has break in it
47
Q

replicative transposition involves

A

transposon duplication in its original position; it is not lost from donor strain

48
Q

Describe conjugation in E. coli

A
  • E. Coli can only be conjugated by a narrow host range of plasmids
49
Q

Describe broad host range plasmids

A

can be conjugated by a large host range of plasmids

50
Q

What is necessary for conjugation?

A

replication origin and machinery recognised

51
Q

Describe some bacteria with a broad range of host plasmids

A
  • Pseudomonas
  • Rhizobium
52
Q

How can DNA integrity be protected from foreign DNA?

A

using various genetic elements such as restriction endonucleases which express methyltransferase, or spacers

53
Q

Give an example of a restriction endonuclease used to protect DNA integrity against foreign DNA

A
  • EchoK
  • EchoR1
  • K9 E. Coli’s
54
Q

Describe the action of methyltransferase in the protection of DNA integrity against foreign DNA

A
  • methylates the recognition sequence of the DNA, to protect it
  • non-methylated DNA will be cut, but methylated DNA will not.
55
Q

Describe spacers, that protect DNA integrity from foreign DNA damage

A
  • past viral DNA that has been integrated
  • adaptive immunity
  • e.g. CRISPR arrays

Hilary Term 1 (61 decks)

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