TAG Theoretical and Applied Genetics 84(7-8): 798-802 (Sept 1992)
Chromosomal and cell size analysis of cold tolerant maize
L. M. McMurphy and A. L. Rayburn
Department of Agronomy, University of Illinois, Turner Hall, 1102 S. Goodwin Ave., 61801 Urbana, IL, USA


C-band number, guard cell length, and chloroplast number per guard cell were determined for eight maize populations. These populations consisted of maize selected for cold tolerance at the University of Nebraska as well as the original unselected populations. The genome size of these populations had previously been determined. C-band number fluctuated concertedly with the changes in genome size indicating that deletions and additions of constitutive heterochromatin occurred during selection, resulting in altered genome sizes. Guard cell size of all the cold tolerant populations was greater than the cell size of the respective nonselected populations. Chloroplast number per guard cell was also higher in all the cold tolerant populations than in their parental populations, but the increases were not statistically significant. The results indicate that changes in genome size that occurred during selection for cold tolerance are the result of changes in amounts of C-band heterochromatin and that the selection process results in an increase in cell size in the cold tolerant populations.


The adaptation of plants to growth in cold environments is associated with several changes including adjustments in guard cell size and nuclear DNA amount. Limin and Fowler (1989) found that reductions in guard cell size accompanied adaptation to severe winter climates in the tribe Triticeae. Grime and Mowforth (1982) investigated the relationship of cell size and genome size to growth in different climates and found that plants which complete their life cycles during winter and early spring tend to have larger cells and genomes than those that mature in warmer months.

Bennett (1976) found a direct positive correlation between genome size and adaptation to growth at cooler northern latitudes of the United States for a number of grasses. An opposite phenomenon has been shown to occur in Zea mays, where DNA content decreases with increasing latitude. This decrease in genome size has been postulated to be an adaptation to growth in cooler, moister regions with shorter growing seasons (Rayburn et al. 1985). These observations led to the evaluation of genome size in populations of maize selected for cold tolerance (McMurphy and Rayburn 1991). While no unequivocal association between cold tolerance and genome size was observed, genome size appeared to be involved in adaptation to cold. Those cold-tolerant lines which exhibited a degree of freeze tolerance increased in genome size from the original populations, while the one population which was cold tolerant but extremely freeze sensitive had a decrease in genome size.

Maize chromosomes often contain heterochromatic knobs (McClintock 1929) that have been observed to be correlated with various environmental parameters. Maize populations from northern latitudes have fewer knobs than populations from southern latitudes (Longley 1938; Brown 1949; Rayburn et al. 1985). DNA content has been positively correlated with C-band (the mitotic equivalent of pachytene knobs) composition in maize (Rayburn et al. 1985). In some maize populations from the southwestern United States, however, variation in DNA content did not appear to be entirely accounted for by C-band positive DNA and supernumerary B-chromosomes (Rayburn and Auger 1990; Porter and Rayburn 1990).

The objective of the study presented here was to determine if cold tolerance in maize is associated with cell size and chloroplast number and whether the genome size variation of the cold-tolerant populations is a result of variation in amount of C-band (knob) heterochromatin.


Selection for cold tolerance resulted in changes in C-band number and guard cell lengths in selected maize populations, and the former changed concertedly with reported changes in genome size (McMurphy and Rayburn 1991) of the selected populations. These facts indicate that all or most of the genome size changes were the result of changes in amount of repetitive heterochromatic DNA. These DNA sequences have no coding function, so the effect of deletion or addition of this genetic material is primarily nucleotypic (Bennett 1971).

The mean cell size of each cold-tolerant population was greater than that of its corresponding unselected population. This finding is in agreement with the hypothesis of Grime and Mowforth (1982) that plants which complete their life cycles during early spring and winter have larger cells than those that mature in the summer. Limin and Fowler (1989) would have predicted cells of the cold-tolerant populations to be smaller than those of the nonselected populations. A critical difference between the two studies is that Grime and Mowforth observed plants that were able to actively grow and reach maturity in cold environments, whereas Limin and Fowler studied the ability of plants to survive severe winter climates so that they could complete their growth when temperatures were more suitable. The selection pressures used in developing the cold-tolerant maize lines were similar to the early spring growth conditions that Grime and Mowforth associated with large cells. Also, Limin and Fowler focused on the tribe Triticeae. It has previously been determined that although a number of grasses, including some of the Triticeae, demonstrate an increase in genome size as latitude of cultivation progresses northward, maize follows a different trend, with genome size decreasing at northern latitudes. This may indicate totally different methods of adaptation to cold between Triticeae and maize.

It is interesting to note that in the case of NA/NA CT, changes in cell size did not reflect changes in genome size. Genome size decreased significantly in NA CT compared to NA, whereas cell size increased significantly in NA CT. Previous studies have indicated a correlation between genome size and cell size (Butterfass I973; Limin and Fowler 1989). Our study suggests that cell size may be more greatly influenced by environmental adaptations than by genome size. DNA content may dictate a range of permissible cell sizes from which an ideal size is selected based on environmental pressures.

Although no statistical differences were found between chloroplast numbers of selected populations versus their nonselected counterparts, there was a trend toward higher chIoroplast numbers in the cold-tolerant populations. This trend follows the changes seen in cell size. These findings, although statistically nonsignificant, support the hypothesis of Pyke and Leech (1987), which states that celI size is more important than genome size for determining chloroplast number per guard cell. The differences in genome size among the populations examined are due to changes in heterochromatin amounts caused by adaptation to growth in cold environments. Populations which are both cold tolerant and freeze tolerant exhibit increased amounts of heterochromatin, whereas decreased heterochromatin amounts accompany cold tolerance alone. Cold tolerance with or without freeze tolerance is associated with increased guard cell size and increased chloroplast number per guard cell.