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In bacterial genetics, some of the most important tools are surprisingly simple. One of them is a piece of velvet stretched over a wooden block. Esther Lederberg used it, with Joshua Lederberg, to answer a question that matters every time a doctor prescribes an antibiotic: do drugs create resistant bacteria, or do they pick out ones that were already there? This post covers who she was, how replica plating works, her discovery of a hidden virus, a worked example you can follow, and what students can take from her work.
Who was Esther Lederberg?
Her dates, 1922 to 2006, appear in the title of her chapter in the reference book Pioneers in Microbiology. Joshua Lederberg's Nobel biography records that they married in 1946 while he was at Yale, that she had earned her MA at Stanford with G. W. Beadle in 1944-1946, and her PhD at the University of Wisconsin in 1950, and that she worked full time as a research associate.
Joshua Lederberg won half of the 1958 Nobel Prize for gene transfer in bacteria, which we cover in our post on his work. Esther did not share the prize. In his Nobel lecture he said his studies had been done with many colleagues, "above all my wife". This post is about her own contributions.
How does replica plating work?
The method was published by Joshua and Esther Lederberg in the Journal of Bacteriology in 1952 as Replica plating and indirect selection of bacterial mutants. Joshua's Nobel lecture describes it in one line: "transposing a pattern of surface growth from plate to plate with a sheet of velvet".
The steps are:
Spread bacteria thinly on an ordinary nutrient plate, the master plate, and let separate colonies grow.
Press sterile velvet gently onto the master plate. The fine fibres pick up a few cells from every colony, in the same positions.
Press the same velvet onto one or more new plates, keeping the orientation. Each new plate gets a copy of the colony pattern.
If a new plate contains a drug, only colonies with resistant cells grow there. Their positions point back to the matching colonies on the master plate.
The master plate never touches the drug. So if a colony there turns out to be resistant, its resistance cannot have been caused by the drug.
A worked example with sample numbers
Here are sample numbers for a classroom version of the experiment.
A master plate has 200 colonies of E. coli, none exposed to streptomycin.
You replica-plate onto a streptomycin plate. Three colonies grow, at positions you mark.
You pick the three matching colonies from the master plate and grow them in plain broth with no drug.
You test these cultures on streptomycin. They grow. Cultures from colonies that failed on the drug plate do not.
What does this show? The three colonies were resistant before they ever met streptomycin, because the cells you tested came from the untouched master plate. The drug only revealed and selected them. Repeat the cycle and you can obtain a pure resistant line that has never been exposed to the drug.
Common mistakes:
Rotating the velvet. If the orientation changes between plates, positions no longer match. Plates are marked so they can be lined up.
Thinking the drug caused the mutation. This is the idea replica plating was used to test.
Confusing replica plating with dilution. Joshua's lecture calls them alternative methods of indirect selection; replica plating is simply more convenient.
What else did Esther Lederberg discover?
The strain E. coli K-12, which the Lederbergs used for crosses, carries a temperate phage, lambda. In the Nobel lecture Joshua says they had expected the prophage to behave as a separate unit, "but Dr. ESTHER LEDERBERG'S first crosses were quite startling in their implication that the prophage segregated as a typical chromosomal marker". The work appeared as Genetic studies of lysogenicity in Escherichia coli in Genetics in 1953, with Esther as first author.
She was also an author on the papers that described F, the factor that lets E. coli act as a donor in conjugation: Sex compatibility in Escherichia coli (1952) and An infective factor controlling sex compatibility in Bacterium coli (1953), with Luca Cavalli-Sforza and Joshua Lederberg. The lecture explains that F can sit in the cytoplasm, in F+ strains, or in the chromosome, in Hfr strains.
What can a student take from her way of working?
Build a method that removes the doubt. Replica plating did not just suggest resistance came first; it made the alternative impossible.
Trust a startling result enough to follow it. Her lambda crosses contradicted what the group expected, and that was why they mattered.
Credit is not the same as contribution. Read author lists, not just prize lists.
If conjugation, transduction and lysogeny keep blurring together, the Fermi Concept Library covers every concept in biology, each with its own lesson and its place in the map. For the earlier bacterial experiment on transformation, read about Frederick Griffith.
Try it now: sketch a master plate with ten colonies and two drug plates, and work out which colonies are resistant to both drugs.
