Genetics Ch.7: Bacterial and Phage Genetics
How many chromosomes do bacteria have
1 circular haploid chromosome
How to score phage phenotypes
1. Coinfect 2. Isolate progeny phage 3. Plate on mixed lawn (few phage)
What characteristics do we look at for bacterial
1. Colony morphology 2. Resistance 3. Auxotrophs
How do you mate phage
1. High concentration of both + small number of bacteria 3. Bacteria are likely to get coinfection 4. Crossover between phages inside host
Arguments for why conjugation isn't caused by mutation
1. Low probability of multiple mutations on the same gene 2. When plated individually, no recombinant
What are the 2 stages of phages
1. Lytic cycle 2. Lysogenic cycle
2 phenotypes for phage genetics
1. Plaque morphology 2. Host strain
Who discovered cis-trans complimentation
Benzer
What does an catabolic pathway for an auxotroph mean
Can't break something down (i.e. can't use galactose as a carbon source)
What is an auxotroph
Can't grow on MV
What does an anabolic pathway for an auxotroph mean
Can't make something
What is the functional definition of the gene
Cistron: smallest unit that shows us cis/trans effect (cis and trans are different)
What did fine structure mapping show us
Colinearity between mutations (on DNA) and proteins - order of mutations matched the order of amino acids changed on its protein
What kind of phage are in generalized transduction
Defective phage
Who worked out the phage cross
Delbrück
What is fine structure mapping
Detecting very rare XO (XO within a gene)
What is mutation mapping
Determining if a mutant is in the same gene or different genes
What is d
Distance in minutes
What happens in generalized transduction
Entire hot genome gets cut into pieces - each daughter phage contains random piece of the host genome
What determines lysogenic or lytic phase
Environmental conditions (bad conditions -> lysogenic phase)
What does F factor code for
Making of sex pilus that connects 2 bacteria together
What is something eukryotes have that prokaryotes dont
Meiosis
What are genes that compliment each other
Mutants that are on different genes (in Cis or Trans) -> produce wildtype
What does failing to compliment mean
Mutations are on the same genes (in trans) - any mutation in the gene makes nonfunctional protein
What does transformation involve
Naked DNA transfer
What would a T2H+ clearing look like
Only kills one strain but not the other (on mixed plate) - turbid plaque
What is the lysogenic cycle
Phage incorporates into host, will lay dormant for some time
What does transduction involve
Phage transfers DNA
What are temperate phages
Phages with 2 stages
Techniques of phage genetics and what it is called
Plate lawn of bacteria, phage will land on it and create a clearing from killing the bacteria called a plaque
What is specialized transduction
Prophage inside host DNA is supposed to cut itself out to get packaged -> sometimes it makes mistake and packages host DNA with it
What are bacteriophage made of
Protein coat surrounded by nucleic acid (RNA or DNA, circular or linear, SS or DS)
Common way to do screen with bacteria and who did it
Replica plating - Lederberg
How does transformation work
Taking DNA up from the environment using special importing protein
What is induction
Temperate phage switches from lysogenic cycle to lytic cycle because of environmental stressers that are going to kill the host first
What does higher cotransduction frequency mean
The closer the genes are together on the chromosome
What is recombination in bacteria and what is not involved
Transfer and XO of DNA - no gametes involved
What is transduction
Transfer of DNA using bacteriophage (phage)
How do Hfr transfer DNA
Transfers host DNA to recipient, F factor is transferred last
Only way to get mutation on the same gene in cis
Very narrow crossover
What was found in terms of likelihood with FSM
XO not completely random - there are hotspots
Cotransduction frequency equation
cd = (1-d/L)^3
How to do fine structure mapping
rII mutants: only grow on strain B. rII+: B or K12. Independently isolated 300 rII mutants and cross them. Coinfect -> progeny -> plate on k12 (only rII+ can grow - selection)
What is the selection example with met- met+ called
suicide selection
What are Hfr strains
High frequency recombinant - F factor is inserted into the chromosome
What is conjugation mapping not good for
High resolution mapping - can get minutes but not seconds
What kind of transfer do bacteria do
Horizontal nonreciprocal transfer (individual to individual)
What is cotransduction
How often both genes are transferred
How would mutations on the same gene be able to compliment
If they are in cis
What causes specialized transduction
Inaccurate cutting of the host - only genes flanking the prophage
Initial discover of transduction
Infect donor cell -> collect daughter phage -> use them to infect recipient cell -> some genes get transferred with the phage (temperate phage mostly)
How do to selection for auxotroph
Inoculate met- and met+ in minimal liquid media with ampicillin -> amp- won't grow so it won't get killed, met+ will try to grow so it will get killed. Then plate all on complete/supplemental media -> only met- will grow because all met+ is dead
How do lytic cycle phage transfer DNA
Insert their N.A into recipient, replicate inside the cell mostly using host machinery, lyse the host, then spread
What does the temperate phage do in the lysogenic cycle with its DNA and what is it called
Inserts its DNA into the hosts chromosome = prophage -> enters dormant phase
How does gene mapping using conjugation work
Interrupted mating: Interrupt mating at different times to see which genes have transferred
Who worked out transduction
Lederberg
Who worked out conjugation
Lederberg and Tatum
What is cotransformation
2 genes transform together - likelihood 2 genes transform together is related to how close they are
What is L typically for phages
2 minutes
Procedure/selection for generalized transduction
2-point cross: Select for 1 gene, score for the other
2 kinds of transduction
1. Specialized transduction 2. Generalized transduction
3 kinds of bacteria genetic transfers
1. Transformation 2. Transduction 3. Conjugation
How does recipient take on transferred host DNA
2 crossovers flanking the genes
What is F factor
A selfish plasmid - mini chromosome
How do select for 1 gene and score for other
After str kills donor (strs) bacteria, take daughter phages from and allow them to infect recipient. Replica plate on media lacking each gene to see which got which genes.
Why is transformation important
For modifying genes, not mapping them
Who discovered transformation
Griffith - avirulent bacteria + heat killed virulent bacteria
How can resistance be different in bacteria
Resistant to a particular drug
Fundamental difference between screen and selection
Selection is killing non-desired phenotypes. Screen you check each colony
What does conjugation involve
Sex pilus
2 types of plaque morphology
Sharp (rapid lysis) or fuzzy edges (slower lysis)
How does F factor transfer
Single strand through the pilus then they both replicate to be DS
What is L
Size of the phage chromosome in minutes
What happened when they plated the two --++ x ++-- on the MV plate
Some ++++ grew at 10^-5