Embryogenesis in tissue cultures of the domestic carrot, Daucus carota l.

Two suspension culture lines derived from seedling roots of Daucus carota L. and which showed contrasting forms of morphogenetic expression rhizogenesis and embryogenesis - have been used to compare these two processes and to attempt to define how far the morphogenetic pathway followed can be modified by the conditions of culture. Embryogenesis was favoured by lowering the shaking speed from the normal rate (120 r.p.m.) to 60 r.p.m., whereas rhizogenesis could be enhanced by selecting the larger clumps in the culture. Quantitative methods have been developed for the estimation of the embryogenic potentials (E.P.) of callus and suspension cultures, and these methods have been used to follow the decline in the capacity for expression of totipotency which occurs with subculturing. Suspension cultures have been grown in a chemostat in an attempt to obtain a system in which growth and embryogenic potential may be controlled and maintained at steady state levels. A number of culture lines have been maintained by serial subculture on different media or under differing cultural conditions, and their rates of loss of E.P. compared; some of these cultures showed a marked decline in absolute E.P. after about 6-8 passages, and an associated increase in sensitivity to inhibition of embryogenesis by 2,4-D. Microscopic observations showed that decline in E.P. was associated with a change in the predominant cell type of the cultures. Plating of cultures of low or declining E.P. yielded some clones of restored E.P. and some clones of nil E.P. indicating that the parent cultures consisted of more than one cell population. The decline in E.P. has been found to be associated with increase in the prominance of autotetraploids, aneuploids, and occasionally of octoploids in the cultures. Under the normal cultural conditions, these had a much lower capacity for embryogenesis than diploid cells; however, it has not been possible to establish that loss of E.P. is entirely accounted for by change in the chromosome complement of the cultured cells. In an attempt to explain the accumulation of polyploid cells in the cultures, an experiment was designed in which equal numbers of cells from a diploid culture of high E.P. and a tetraploid culture of low E.P. were mixed together. It was found that tetraploid cells became predominant in the mixed culture within three passages, and E.P. declined. The tetraploid culture utilized sucrose more rapidly than the diploid culture and showed a greater dry weight increase during the exponential phase of the growth cycle. Thus, it was concluded that loss of E.P. in serially r subcultured tissue occurred because a non-embryogenic population of cells competed successfully with an embryogenic population and so came to form the predominant cell type. Techniques have been developed for the raising of mature plants from the embryos obtained from carrot tissue cultures; field tests have indicated that there is greater variation amongst the progeny of plants raised from tissue cultures as compared with plants of the standard breeding line. Finally, problems associated with the use of embryogenesis in tissue and cell cultures for micropropagation are discussed.