Transduction Flashcards
Generalized transduction overview steps
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)
Specialized transduction overview steps
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)
transduction is the transfer of bacterial DNA from one cell to another by ________
a bacteriophage (transducing phage)
two types of transductions & their defintions
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
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 ______
-DNA from host
-anywhere on chromosome
-recombinants/transductants
all generalized transducing phages share 3 features:
1) non-specific pac sites
2) phage DNA is packaged using a headful mechanism
3) broad host range
T7: pac site is the DNA sequence where ______. They are often found in _______ of phage DNA produced during ________ replication.
-phage DNA is initially cut for packaging into phage heads (T7)
-concatemers
-rolling-circle
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.
-headful packaging
-bacterial chromosome
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
-paclike
-phage chromosome
most useful transducing phages have a ____ host range for adsorption. transducing phage do not have to be able to _______, only attach & inject DNA
-broad
-multiply in recipient
Best known generalized transducing phages
P1 (E.coli)
P22 (S. tryphimurium)
P1 phage length of DNA packaged? host DNA is packaged from _____?
-100 kb
-ends
P22 phage length of DNA packaged? host DNA is packaged from _____?
-44 kb
-pac-like sequences
Two main uses of generalized transduction
1) Mapping
2) Strain construction
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.
-50-100
-cotransduced
-greater
mapping with generalized transduction: gen. transduction is very ____. To detect transduction/measure co-tranduction, transductants are plated on media where _____
-rare
-where only recombinants grow
mapping with gen. transduction: if 2 markers are linked = _______; if 2 markers unlinked = ______
-cotransduction
-no cotransduction
cotransductant is the _____ species
minor
frequency of co-transduction is _____ proportional to distance between genes
-inversely
Co-transduction frequency = _______. can use co-transduction frequencies to construct ______
-# cotransductants/total transductants X 100
-genetic maps
mapping with gen. transduction technique
-one marker is selected for
- presence of other markers screened for
-calculation of co-transduction frequencies allows construction of genetic map
Flipping DNA ______ does not change gene order.
-horizontally
To definitively determine gene order with gen. transduction –> ______
-3-FACTOR CROSS
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)
- # of recombination events/cross-overs-2 recombination events
-even
-odd
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 ____
-two
-the correct order
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. ____
-strains that differ only in gene being studied
-antibiotic resistance
Specialized/Restricted Transduction: bacteria DNA is transferred between donor and recipient as part of a _______ that arises from aberrant excision of _______
-specialized transuding particle
-integrated prophage
what is a specialized transuding particle?
phage & bacterial DNA linked in continuous molecule and packaged in same phage head, contains only regions of DNA that flank prophage
what are the steps of formation of specialized transducing phage?
1)prophage induction (return to lytic growth)
2) rare mistake in excision occurs
3)bacterial DNA adjacent to prophage is excised with phage DNA
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 _____
-aberrant excision occurs
-required
-defective
specialized transducing particle are very rare -> _____ excision events happen at low rate.
-erroneous
once specialized transducing phage integrated into chromosome of recipient, requires _______ to produce more phage particles; it provides missing phage functions
-helper phage
You create _____ carrying specialized transducing phage and wild-type helper phage to permit isolation of high frequency transduction (HFT) lysates
-di-lysogens
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
-virulence factors or toxins
-morons
-promoters
-elevated
how does lysogenic conversion of lambda-like phage happen? -EHEC (enterohemorrhagic E.coli)
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)
how does lysogenic conversion happen in Vibrio cholerae?
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.
