CHROMOSOME DUPLICATION AND GENETIC RECOMBINATION 189 



solvent of appropriate density, approximately IM cesium chloride in 

 water. The small molecules of cesium chloride are distributed across the 

 cell of the ultracentrifuge in a continuous density gradient, but the huge 

 DNA molecules collect in a narrow band corresponding to their density. 

 Molecules of uniform density are distributed in the region of the band 

 as a result of thermal agitation, and from the band width the molecular 

 weight can be computed. If there is a distribution of density differences 

 among molecules of a particular sample, there will be a further broad- 

 ening of the band. If the molecular weight and size homogeneity of the 

 sample are known from other experiments, then the bandwidth can be 

 used to compute density differences within the sample. This method is 

 so sensitive that N^'^DNA and N^^DNA separate readily into distinct 

 bands. 



The experiment consists of growing bacteria (£. coli) with an N - 

 containing nitrogen source until the DNA is well labeled, then allowing 

 anv desired amount of growth with an N ^^ nitrogen source, and finally 

 observing the pattern of DNA distribution in the ultracentrifuge. 



The results are remarkably clear cut, as can be seen in Figure 7.4. 

 Initially only one band is found, corresponding to N^^DNA. After 

 one doubling in the presence of N^^, again one band is found, but in a 

 new position. It is intermediate between N^^DNA and N^^DNA. 

 After a second doubling, two bands are found of equal DNA concen- 

 tration. One is the intermediate band, and the other is still less dense, 

 and is in the position of N '^DNA. Subsequently, the intermediate band 

 decreases by half after each doubling and the N^^DNA band increases 

 correspondingly. Similar results have been obtained with Chlamy- 

 domonas and with mammalian cells in tissue culture. 



The intermediate band contains equal amounts of N ^^ and N^°DNA, 

 combined in such a way that neither the salt concentration nor the 

 centrifugal forces dissociate them. However, by heating under condi- 

 tions that break hydrogen bonds, the mixture has been dissociated into 

 two subunits, one containing all N^^ and the other all N^'^, and each 

 with a molecular weight half that of the original molecule. It may be 

 inferred, therefore, that the band appearing after one doubling contains 

 hybrid molecules-, each consisting half of an N ^^ subunit and half of an 

 N^^ subunit. It is probable that the subunits are the two strands of 

 the double helix (although more complicated models can be devised to 

 account for the data). 



On the hypothesis that the subunits are individual polynucleotide 

 strands, the interpretation of the experimental results is straightforward, 

 as diagrammed in Figure 7.3. In the first replication, each N ^^ strand 



