Lecture 7 Flashcards
(34 cards)
What is a bacteriophage?
Viruses that parasitize bacteria
What makes bacteria a good model organism?
They grow quickly (fast dividing) in huge number, take up little space, and allow us to see rare events
How does heredity work in bacteria?
Fundamentally the same as eukaryotes: DNA codes for proteins via RNA, a copy of DNA is passed on to the daughter cell.
- Haploid
- Reproduce asexually
- May exchange DNA with other organisms by accident through transformation, transduction, or conjugation
- Evolution is through mutation and other “accidents”
What is spread plating?
A stock of bacterial cells can be grown in liquid medium containing nutrients, then a small number of bacteria from the liquid suspension can be spread on a solid agar medium. Each cell will give rise to a colony of cells that have the same genotype and phenotype. When the cells reach more than 10^7 they are visible as a colony.
What is a prototroph?
Wild type bacteria that can grow on minimal medium (inorganic salts, carbon source for energy, and water). It has the functional enzymes necessary to make all needed macromolecules.
What is a auxotroph?
Mutant bacteria from prototroph, they are unable to grow on minimal medium. They require supplementation with product the mutant cannot synthesize for itself in order to grow.
What are resistant mutants?
Mutant bacteria that can divide and form colonies in the presence of an inhibitor whereas the wild type bacteria would be susceptible to the inhibitor.
What is transformation?
Direct uptake of DNA from the environment generally from dead or lysed cells. Unless this DNA has a origin of replication (ori), it must recombine into the chromosome. The transforming DNA is incorporated into the bacterial chromosome by a process analogous to the double-recombination events observed in Hfr × F−
crosses.
How can double transformation be used for mapping genes?
Genes located close together may be carried on the same piece of transforming DNA. When the DNA is taken up it causes double transformation. If genes are linked, then the proportion of double transformants will be greater than the product of single-transformant frequencies.
How was conjugation discovered?
Lederberg and Tatum demonstrated that genetic recombination between bacterial genotypes is possible. The basic concept: two auxotrophic cultures (A− and B−) are mixed, yielding prototrophic wild types (WT). Cells of type A− or type B− cannot grow on an unsupplemented minimal medium because A− and B− each carry mutations that cause the inability to synthesize constituents needed for cell growth. When A− and B− are mixed for a few hours and then plated, however, a few colonies appear on the agar plate. These colonies derive from single cells in which genetic material has been exchanged; they are therefore capable of synthesizing all the required constituents of metabolism.
What is conjugation?
Formation of pilus between donor cell containing sex factor and recipient cell. Donor replicates its plasmid and donates copy to recipient. Transfer is unidirectional. Plasmid contains sex factor. If F factor is integrated into chromosome, the entire chromosome will attempt to transferred to the recipient.
What is cross-feeding? U-tube experiment?
Cross-feeding is the idea that strains don’t actually exchange genes but instead leak substances that the other cells can absorb and use to grow. It was disproved by the U-tube experiment where filter separated A- and B- mutants so that no contact was possible but diffusion of molecules was. After incubation and plating, no protrophic cells grew. This proved that cell to cell contact is necessary.
What did Hayes discover how about the sharing of genetic material?
Sharing is unequal. One strain donates it alleles to convert the other. Donors are F+ which contain the fertility factor. F- are converted to F+ and can then pass on the fertility factor themselves. F+ is immune to superinfection (can’t put a second F plasmid into F+).
What are some important properties of the F factor?
F factor is a plasmid that can replicate in the cytoplasm independent of the host chromosome.
- Directs the synthesis of sex pilus
- Codes for pilus proteins, proteins that mobilize plasmid into new cells
- Has genes that inhibit superinfection
- contains oriV for plasmid replication and oriT for conjugative transfer
- F+ has pili that stick out from surface, when a pilus contacts a F- cell, it contracts and proteins facilitate fusion of membranes
How is the F factor transferred?
Rolling circle replication: A single-stranded copy of plasmid DNA is produced in the donor cell and then passes into the recipient bacterium, where the single strand, serving as a template, is converted into the double-stranded helix.
What is an Hfr cell?
Integration of the F factor into the bacterium’s chromosome. This integration may occur due to homologous recombination between the F factor and chromosome. The region of homology are segments of transposable elements known as the insertion sequence.
What are some important properties of Hfr cells?
- Still make pili
- Will try to transfer entire chromosome into recipient cell which would take about 100min to complete
- Can put genes from chromosome into F- cell as non-replicating linear DNA
- Genes may recombine into chromosome of F-
What is interrupted mating? What is it used for?
Hfr cells and F- cells are mixed together then placed in a blender to separate mating cell pairs (interrupted mating). The samples are then placed on medium to kill donor Hfr cells. Surviving cells were tested for presence of alleles from donor Hfr genome.
Describe linear transmission of Hfr genes to a F- cell.
- Each Hfr strain contains F at a specific place in the chromosome
- Transfer begins from a fixed point in the middle of F
- The orientation of F determine which genes transferred first
- Part of F (oriT) transferred first but remaining part will be the very last
- Recipient will not become F+ unless entire chromosome transferred (rare)
- Donor alleles appear in a specific sequence
- Single stranded copy is first transferred then converted into double helix. Double cross over may occur and insert donor DNA.
What are the two states the fertility factor can exist in?
The plasmid state as a free cytoplasmic element o r the integrated state as a continuous part of a circular chromosome.
What are two ways to map bacterial chromosomes?
- Broad scale chromosome mapping by using time of entry
2. Fine scale chromosome mapping by using recombinant frequency
What is broad scale mapping by using time of entry?
Constructing linkage maps from interrupted mating results. Units of map distance are in minutes.
What is fine scale mapping by recombinant frequency?
Examine recombination frequency after interrupted mating. Where crossovers occur determines which genes are incorporated. Recombinant frequency of events are used to obtain relative sizes of segments.
What is the endogenote?
Complete genome from F- cell
