Microbial Recombination and Gene Transfer Questions and Answers

Answer: B

Correct answer: a DNA sequence that can move to a new position within the genome

A transposon, or transposable element, is a DNA sequence able to change its position within the genome. They were discovered in maize by Barbara McClintock in the 1940s, work for which she received the Nobel Prize in 1983 - long after it was first dismissed. The simplest form is the insertion sequence, carrying only the transposase gene needed for its own movement, flanked by inverted repeats. A composite transposon has two insertion sequences flanking additional genes, which are carried along with it - and those genes are frequently antibiotic resistance determinants, which is how resistance moves between plasmids and chromosomes. Movement may be conservative, cutting the element out and pasting it elsewhere, or replicative, copying it so that both sites end up occupied. Insertion into a gene disrupts it, so transposons cause mutations, and that property is exploited deliberately in transposon mutagenesis to identify gene function.

Answer: D

Correct answer: their order along the donor chromosome

In an interrupted mating experiment an Hfr donor is mated with an F− recipient and the pairs are separated at intervals - classically in a kitchen blender. Testing which markers have arrived at each time reveals the order of the genes along the donor chromosome. The method works because Hfr transfer is sequential: it begins at the point where the F plasmid integrated and proceeds linearly, so a gene nearer that origin enters earlier. The resulting map is measured in minutes rather than in recombination frequency, and the whole E. coli chromosome takes about 100 minutes to transfer - which is why the standard genetic map of E. coli is a circle of 100 minutes. Different Hfr strains have the plasmid integrated at different sites and in either orientation, so each transfers a different starting point and direction. Combining several such maps was what established that the chromosome is circular. The technique is due to Wollman and Jacob in the 1950s.

Answer: B

Correct answer: Site-specific recombination

Site-specific recombination requires no general homology. It acts at short, defined sequences recognised by a dedicated integrase, which is exactly how the lambda phage genome enters the bacterial chromosome: the phage attP site and the bacterial attB site share only a short core sequence, and integrase catalyses the exchange between them.

General recombination, by contrast, depends on extensive homology and RecA. Replicative recombination is the mechanism of transposition, and mutagenic recombination is not the standard name for phage integration.

Answer: D

Correct answer: Lederberg and Tatum

Joshua Lederberg and Edward Tatum demonstrated genetic recombination between bacterial cells in 1946. Mixing two multiply auxotrophic strains of Escherichia coli K-12, they recovered prototrophs at a frequency far above the reversion rate of either parent, showing that genetic material had been exchanged. Davis's U-tube experiment later established that cell contact was required, defining conjugation.

Ochoa and Kornberg were recognised for the enzymatic synthesis of RNA and DNA, Khorana for work on the genetic code and oligonucleotide synthesis, and Muller for showing that X-rays induce mutations.

Answer: B

Correct answer: size, with smaller fragments migrating further

Gel electrophoresis separates DNA fragments by size. The DNA is loaded into wells in an agarose gel and an electric field applied; because the phosphate backbone gives DNA a uniform negative charge per unit length, all fragments migrate towards the anode at a rate governed by how easily they thread through the pores of the gel. Smaller fragments travel further. The uniform charge is what makes the method work: since charge is proportional to length, charge-to-mass ratio is essentially the same for every fragment, and only sieving by the gel matrix distinguishes them. Fragments are visualised by staining with ethidium bromide or a safer alternative and viewing under ultraviolet light, and their sizes are read against a ladder of known standards run alongside. Agarose suits fragments from a few hundred bases to tens of kilobases; polyacrylamide resolves smaller ones more finely. Very large fragments need pulsed-field electrophoresis, which periodically changes the field direction.

Answer: A

Correct answer: transformation

Transformation is the uptake of naked DNA from the environment by a competent bacterium and its incorporation into the genome, usually by homologous recombination. It was the first mechanism of bacterial gene transfer discovered, in Griffith's 1928 pneumococcal experiment, and the one Avery, MacLeod and McCarty used in 1944 to identify DNA as the genetic material.

Conjugation requires cell-to-cell contact and transduction requires a phage vector. Lysogenic conversion is the change of phenotype caused by genes carried on a resident prophage, which is a consequence of lysogeny rather than a route of DNA uptake.

Answer: D

Correct answer: transduction

Penicillin resistance in staphylococci is classically acquired by transduction. The penicillinase gene lies on a plasmid, and staphylococcal phages package that plasmid and carry it into new cells, which is the principal route by which the resistance spread through hospital strains.

Staphylococci have no efficient natural conjugation or transformation system comparable with that of the enteric bacteria, so those routes are not the usual explanation. Mutation can alter a chromosomal target, but the beta-lactamase responsible here is an acquired plasmid-borne enzyme rather than a mutated host gene.

Answer: D

Correct answer: transduction

Penicillin resistance in staphylococci is classically acquired by transduction. The penicillinase gene lies on a plasmid, and staphylococcal phages package that plasmid and carry it into new cells, which is the principal route by which the resistance spread through hospital strains.

Staphylococci have no efficient natural conjugation or transformation system comparable with that of the enteric bacteria, so those routes are not the usual explanation. Mutation can alter a chromosomal target, but the beta-lactamase responsible here is an acquired plasmid-borne enzyme rather than a mutated host gene.

Answer: B

Correct answer: A cluster of genes transcribed together from a single promoter

An operon is a group of adjacent genes transcribed together as one messenger RNA from a single promoter, so that functionally related proteins are produced in coordinated amounts. The arrangement is characteristic of prokaryotes and was described by Jacob and Monod in 1961. Its parts are the promoter, where RNA polymerase binds; the operator, where a regulatory protein binds; and the structural genes. A separate regulator gene encodes the protein that binds the operator. The lac operon is inducible: a repressor blocks transcription until lactose - through its isomer allolactose - binds the repressor and removes it, so the genes are expressed only when their substrate is present. The trp operon is repressible: it is transcribed until tryptophan accumulates and activates the repressor, switching it off. The two illustrate the general principle that catabolic pathways are induced by their substrate and anabolic ones repressed by their product.

Answer: A

Correct answer: Col plasmids

Col plasmids encode colicins - bacteriocins that a bacterium secretes to kill closely related strains competing for the same niche. Colicins act by forming pores in the target's membrane or by cleaving its nucleic acids, and the producing cell protects itself with an immunity protein encoded on the same plasmid. Col plasmids are typically small and present in multiple copies.

R plasmids carry drug resistance determinants, degradative plasmids encode enzymes for unusual substrates such as toluene or camphor, and virulence plasmids carry determinants that make the host pathogenic to a higher organism. Only the Col plasmid is aimed at other bacteria.