CELLULAR DIFFERENTLYTION IN THE SLIME MOLD 235 



modified scheme which assumed occasional syngamy and meiosis, only 

 coincidental with and not causally related to the fruiting process, and 

 perhaps requiring special conditions of cultivation (as is the case for 

 a wide variety of fungi). These suppositions, too, have met with criti- 

 cism ( Bonner, 1959; Shaffer, 1958 ) . Nevertheless, the cytological data, 

 admittedly incomplete and to some extent equivocal, cannot be gainsaid 

 simply by syllogistic pronouncements; they demand further elucidation, 

 particularly at the genetic level. 



Our own involvement in this problem at first centered upon at- 

 tempts to test whether or not the morphogenetic sequence was coupled 

 mandatorily to a sexual cycle, and the answer appeared to be that it 

 was not. In the course of these investigations, a systematic search was 

 carried out for evidence of genetic recombination. The mutants that 

 were employed differed from the wild type by fruit morphology or 

 pigmentation or by the possession of morphogenetic deficiencies ( i.e., 

 aggregateless, fruitless ) . Unfortunately the primitive state of nutritional 

 control precluded the selection of adequate biochemically deficient 

 stocks, and initial attempts to obtain drug-resistant mutants yielded 

 only unstable dauermodifikations. Thus- it was impossible to select for 

 recombinants by methods used so successfully in Neurospora, E. coli, 

 and other Protista, and examination of clones by random population 

 sampling remained as the only feasible method. The mutant stocks 

 were grown together or allowed to form mixed fruiting bodies under 

 a variety of environmental conditions. The clones obtained from the 

 samples of these mixtures were invariably of either parental type and 

 displayed no sign of recombination. It was concluded that if sexuality 

 did operate, zygotes and recombinants must be exceedingly rare ( <10'^ 

 cells), or there must be mating types which we did not possess, or all 

 our genetic markers were infertile aberrations ( Sussman, 1956 ) . 



The discovery of the anomalous I-cell clones provided the impetus 

 to make another attempt at finding genetic recombination. The rapid 

 shift between the haplo-phase and diplo-phase in these clones made it 

 likely that if sexuality were operative as a rare event, zygotes and re- 

 combinants would be encountered at higher frequency here than in 

 the total myxamoeboid population. A precedent is found in strains 

 (Hfr) of Escherichia coli which display a high frequency of genetic 

 recombination and are derived from the low-frequency (F+) stocks. 

 Accordingly, anomalous clones were obtained from mutant strains by 

 micromanipulation of I-cells in order to employ them as parents in 

 crossing attempts. The results of the initial experiment will be de- 

 scribed. 



Mutant stock br-1 ( brown ) aggregates and fruits normally, and the 

 spore masses accumulate the normal yellow pigment. However, it pro- 

 duces an additional, soluble, red-brown pigment which deeply colors 



