Patterns of variability in DNA content and nuclear volume in regenerating cultures of Nicotiana tabacum

The frequency distributions of DNA content per nucleus were examined in five isonicotinic acid hydrazide (INH) resistant callus lines of tobacco and in developing buds and shoots regenerating from the lines. They were compared with the distribution for a diploid plant, with an estimated 2C value of 7.8 pg. The total range of all the callus cultures was between 5 and 40 pg, with modes between 7 and 16 pg. In the oldest (6 years) callus culture, the mean and standard deviation were lower in the buds developing from the culture than in the undifferentiated callus: however, older shoots again showed a higher mean and more disperse distribution. In contrast, for 4-year-old secondary callus cultures from plants simultaneously regenerated from a second INH-resistant (I 24) line, the mean DNA content per nucleus of the developing buds was higher than that of the undifferentiated callus. Less variation and a more nearly diploid distribution were observed in a 2-year-old callus culture of a fertile plant chosen from the progeny of one of these I 24 plants and in the buds and older shoots regenerated from this culture. Thus, in these five moderately heteroploid cultures a consistent pattern of selection for euploid levels of DNA did not occur during the observed stages of regeneration. Examination of volume as well as DNA content of nuclei in the different tissues showed that variability of nuclear volume measurements, as indicated by coefficients of variation, was correlated with variability of DNA content per nucleus, although measurements of volume and DNA content per individual nucleus were not always highly correlated. It is suggested that high coefficient of variation of nuclear volume, in conjunction with large nuclear size, could serve as a rapid preliminary indicator of highly heterogeneous DNA levels in the nuclear population of Nicotiana cultures.