462 GENETICS OF SOMATIC CELLS 



its frequency and its relation to extrinsic and intrinsic factors. Critical experiments 

 are yet to be done with cells derived from homozygous animals and with close passage- 

 to-passage coordination odn-vitro cultivation and retransplantation to animals isogenic 

 with the original donor. Isogenicity can be best insured by using highly inbred mouse 

 strains, critically tested with skin grafts, and restricting the time elapsing between the 

 origin of the line and its actual testing to the possible minimum, or, alternatively, by 

 using some such technique as frozen storage of spermatozoa and artificial insemination 

 to reproduce the histocompatibility factor equipment of the original donor or animals 

 isogenic with it. 



The possibilities for working out methods for transferring genetic information 

 between somatic cells such as DNA-transformation or virus-mediated transduction 

 have been discussed. A priori the feasibility of some such procedure is no less probable 

 than it had been with bacteria. 



DISCUSSION 



Dr. Burdette: Dr. Leonard A. Herzenberg will open the discussion of Dr. Klein's 

 paper. 



Dr. Herzenberg : The exciting reports of Drs. George and Eva Klein and their 

 collaborators on isoantigenic variations in tumors stimulated us to explore the possibility 

 of extending their genetic analysis in vivo to the cell-culture situation in vitro. In this 

 symposium on methodology, let me indicate some of the methodologic advantages for 

 detailed genetic analysis that cell-culture systems afford us. 



First and foremost, since the demonstrations of Puck, 1029 cloning is a rapid and 

 reproducible procedure in work utilizing cultures of cells. Populations derived from 

 one cell are routinely available, and all the tricks of the microbiologist are applicable 

 to the mammalian-cell system. 



Cells have been cultivated from various tissues of a number of mammals in semi- 

 defined media; 313 that is, in synthetic media to which are added a small percentage of 

 dialyzed serum proteins. These media may contain traces of unknown materials, but 

 the macroconstituents — amino acids, carbohydrates, purines, pyrimidines, most 

 vitamins, and salts — are present in known concentrations. Thus, variations in 

 nutritional needs or susceptibility to metabolic poisons can be searched for, and, if 

 suitable ones are found, these can be employed as cellular genetic markers. One or 

 two nutritional variants have been described. 244 However, these have not yet been 

 too useful in genetic studies due to the difficulty in effectively selecting for these variants. 



A number of drug-resistant variants of mammalian cells in culture have now been 

 found. With some drugs, for example, 6-mercaptopurine, 8-azaguanine, and ame- 

 thopterin, a resistant cell can easily be selected from a large population of sensitive 

 cells, 366, 787 and it has been possible to answer some basic questions about the genetics 

 of resistance to these compounds. With other drugs, for example the fluorinated 

 pyrimidines, efficient selection of individual resistant cells from a background of 

 sensitive cells has proved to be more difficult. Nevertheless, workers in a number of 



