Newcomb's respreading experiment on the spontaneous origin of Ton s to Ton r mutations
Initial number of bacteria per plate
5.1 X 10 4
Hrs. incubation before spraying with T1
5
Bacteria per plate at time of spraying
2.6 X 10 8
Number of cells in microcolony
5 X 10 3
Ton r colonies on parallel plates
undisturbed
5
3
4
8
2
6
total = 28
 respread
  194
14
16
13
13
112
total = 353

8. Newcombe's respreading experiment: This is a simpler experimental version of the ideas explored in Luria and Delbruck's fluctuation experiment.

Newcombe spread 5.1 x 10 4 Ton s on 12 different agar plates and let each cell grow into a microcolony of approximately 5000 cells. 6 plates were left undisturbed and the other 6 plates were respread. Thus in the absense of any T1 phage, the 5000 cells of the 5.1 x 10 4 colonies could have given rise to microcolonies - one for each of the 5.1 x 10 4  X  5000  =  2.6  X  10 8 cells. However immediately after respreading, all 12 plates were sprayed with an excess of T1 to reveal the number of Ton r cells / colonies. The undisturbed plates gave a total of 28 Ton r colonies, in contrast to the much larger 353 Ton r colonies on the 6 respread plates.

It is apparent that at the time of respreading some of the microcolonies were already composed of Ton r cells. For example the 194 Ton r colonies on plate 1 must have originated in one or more of the 5.1 x 10 4 colonies present at the time of spraying - again in support of the prior-mutation hypothesis.

How would you calculate the mutation rate in Newcombe's experiment?

Answer: We use the data from the undisturbed plates and assume that any resistant colony results from a single mutational event. Total number of bacterial divisions at the time of T1 spraying: 6  X  (2.6  X  10 8 - 5.1  X  10 4)

Total number of mutations: 28

Mutation rate: 28 / 6  X  (2.6  X  10 8 - 5.1  X  10 4 ) = 1.8  X  10 -8

Nobel Prize: 1969 For a delighful account of the personal idealism of practitioners of the Arts and Sciences, read Artist vs Scientist in Delbruck's Nobel speech.

Lederbergs' Replica Plating Experiment: A direct, simple and non-statistical, test of the spontaneous prior-mutation hypothesis was performed by Joshua and Ester Lederberg in 1952. When a velvet surface is pressed onto an agar surface containing bacterial colonies, some of the cells from each colony will be picked up by the velvet. Colony cells can then be transferred to a number of fresh plates in succession. When 10 7 colonies were transferred and then sprayed with T1, all daughter plates showed the same spatial pattern of Ton r colonies. If the induction hypothesis was correct one would expect that each plate would have a different number and spatial pattern of Ton r . Otherwise we have to assume that after transfer ( when the plates were subjected to T1) only the same four colony cells were capable of being induced to Ton r by T1. This seems very unlikely and requires ad hoc assumptions to added to the original hypothesis. In contrast the prior-mutation hypothesis explains the identical spatial distribution by virtue of the prior random mutation to Ton r in cells of four colonies which were later transferred to new media.

Click on the cell lineage image for an alternative treatment of the Luria / Delbruck story