Transduction

Question 1

Generalized transduction overview steps

Answer 1

1) Phage infects bacterial cell
2) Host DNA is hydrolyzed; phage DNA and proteins are made
3) Bacterial DNA fragment is packaged in phage capsid
4) Phage infects new bacteria (crossing over/recombination)
5) recombinant bacteria made (different genotypes than donor/recipient)

Question 2

Specialized transduction overview steps

Answer 2

1) Bacteria has prophage integrated between genes A & B
2) Sometimes prophage DNA exits taking the nearby DNA with it
3) Phage particles carry some bacterial DNA with phage DNA
4) Phage infects new bacteria (crossing over/recombination)
5) recombinant bacteria made (different genotypes than donor/recipient)

Question 3

transduction is the transfer of bacterial DNA from one cell to another by ________

Answer 3

a bacteriophage (transducing phage)

Question 4

two types of transductions & their defintions

Answer 4

1) generalized: any region of bacterial DNA can be transferred from bacterium to bacterium
2) specialized: only certain genes close to attachment site of prophage can be transferred

Question 4

generalized transducing phage produce some phage particles that contain only _____. host DNA can be from _____ (generalized). In subsequent infections, host DNA is injected in another bacterium where it can recombine with recipient chromosome to produce ______

Answer 4

-DNA from host
-anywhere on chromosome
-recombinants/transductants

Question 4

all generalized transducing phages share 3 features:

Answer 4

1) non-specific pac sites
2) phage DNA is packaged using a headful mechanism
3) broad host range

Question 5

T7: pac site is the DNA sequence where ______. They are often found in _______ of phage DNA produced during ________ replication.

Answer 5

-phage DNA is initially cut for packaging into phage heads (T7)
-concatemers
-rolling-circle

Question 6

enzyme initially recognizes pac site in concatemers and then ______ enzyme makes regular cuts at unit lengths. similar sites may be recognized in _______ & then bacterial chromosome will be packaged into phage heads.

Answer 6

-headful packaging
-bacterial chromosome

Question 7

generalized transduction: once packaging is initiated at a _____ site, DNA is cut in unit lengths that fill a phage head. so once packaging of bacterial
chromosome starts, ______ unit lengths produced & packaged

Answer 7

-paclike
-phage chromosome

Question 8

most useful transducing phages have a ____ host range for adsorption. transducing phage do not have to be able to _______, only attach & inject DNA

Answer 8

-broad
-multiply in recipient

Question 9

Best known generalized transducing phages

Answer 9

P1 (E.coli)
P22 (S. tryphimurium)

Question 10

P1 phage length of DNA packaged? host DNA is packaged from _____?

Answer 10

-100 kb
-ends

Question 11

P22 phage length of DNA packaged? host DNA is packaged from _____?

Answer 11

-44 kb
-pac-like sequences

Question 12

Two main uses of generalized transduction

Answer 12

1) Mapping
2) Strain construction

Question 13

Two factor cross: each transducing particle carries between ____ genes. genes will only be _____ if they are close enough to be carried on same DNA fragment. the closer genes are to each other the ______ the probability that they will be co-transduced.

Answer 13

-50-100
-cotransduced
-greater

Question 14

mapping with generalized transduction: gen. transduction is very ____. To detect transduction/measure co-tranduction, transductants are plated on media where _____

Answer 14

-rare
-where only recombinants grow

Question 15

mapping with gen. transduction: if 2 markers are linked = _______; if 2 markers unlinked = ______

Answer 15

-cotransduction
-no cotransduction

Question 16

cotransductant is the _____ species

Answer 16

minor

Question 17

frequency of co-transduction is _____ proportional to distance between genes

Answer 17

-inversely

Question 18

Co-transduction frequency = _______. can use co-transduction frequencies to construct ______

Answer 18

-# cotransductants/total transductants X 100
-genetic maps

Question 19

mapping with gen. transduction technique

Answer 19

-one marker is selected for
- presence of other markers screened for
-calculation of co-transduction frequencies allows construction of genetic map

Question 20

Flipping DNA ______ does not change gene order.

Answer 20

-horizontally

Question 21

To definitively determine gene order with gen. transduction –> ______

Answer 21

-3-FACTOR CROSS

Question 22

3-factor cross: analysis based on _____required to generate certain classes of recombinants/ transductants. in transductional cross: minimum of _______ required to integrate short linear DNA into bacterial chromosome. viable recombinants MUST have originated from ____ number recombination events (____numbers will break chromosome)

Answer 22

• # of recombination events/cross-overs-2 recombination events
  -even
  -odd

Question 23

Recombinant types that require more than _____ recombination events will be very rare. Order that produces rare recombinant type (has the most cross over aka 4) will be ____

Answer 23

-two
-the correct order

Question 24

strain construction is used to study gene function. It is essential to compare isogenic strains aka ______. can accomplish this by using transduction or transformation to move mutations into a strain. § can be done even if no way to select -> use closely linked marker, ie. ____

Answer 24

-strains that differ only in gene being studied
-antibiotic resistance

Question 25

Specialized/Restricted Transduction: bacteria DNA is transferred between donor and recipient as part of a _______ that arises from aberrant excision of _______

Answer 25

-specialized transuding particle
-integrated prophage

Question 26

what is a specialized transuding particle?

Answer 26

phage & bacterial DNA linked in continuous molecule and packaged in same phage head, contains only regions of DNA that flank prophage

Question 27

what are the steps of formation of specialized transducing phage?

Answer 27

1)prophage induction (return to lytic growth)
2) rare mistake in excision occurs
3)bacterial DNA adjacent to prophage is excised with phage DNA

Question 28

specialized transducing particle: some phage genes are lost in process depending where _____. phages particles first produced from og excision event -> all ___ phage genes present. phage particles are _____

Answer 28

-aberrant excision occurs
-required
-defective

Question 29

specialized transducing particle are very rare -> _____ excision events happen at low rate.

Answer 29

-erroneous

Question 30

once specialized transducing phage integrated into chromosome of recipient, requires _______ to produce more phage particles; it provides missing phage functions

Answer 30

-helper phage

Question 31

You create _____ carrying specialized transducing phage and wild-type helper phage to permit isolation of high frequency transduction (HFT) lysates

Answer 31

-di-lysogens

Question 32

many prophages carry genes for ______ in pathogenic bacteria. these genes are called ____ (more DNA). not found in all phages. They are expressed from own _____ which are active inside hosts. Expression is ____ by prophage induction

Answer 32

-virulence factors or toxins
-morons
-promoters
-elevated

Question 33

how does lysogenic conversion of lambda-like phage happen? -EHEC (enterohemorrhagic E.coli)

Answer 33

1)prophage induction are repressed by cI protein, cI represses lytic genes until cleaved by RecA (SOS response)
2)SOS response induced in host
3) leads to inactivation of Stx phage repressor of lytic genes
4) elevated expression of stxAB genes
5)cell eventually lysed
6) releases Shiga toxin = Can lead to “Hamburger disease” (HUS)

Question 34

how does lysogenic conversion happen in Vibrio cholerae?

Answer 34

1) Toxin Co-regulated Pilus (TCP) is required for Vibrio’s attachment to intestinal epithelium (TCP is acquired from a filamentous phage).
2) TCP is also receptor for CTXf phage, which carries CTX (cholera toxin) gene.
3)CTX phage infects Vibrio with TCP expressed, bringing in CTX gene, and cell subsequently produces CTX to cause disease.