Plant Physiol. 1956 Sep;
VERNALIZATION IN PEAS1
H. R. HIGHKIN2
EARHART PLANT RESEARCH LABORATORY, DIVISION OF BIOLOGY,
CALIFORNIA INSTITUTE OF TECHNOLOGY, PASADENA, CALIFORNIA
manuscript received June 5, 1956.
2 Report of work supported by a grant from the National Science Foundation.
A great number of plants have been shown to require a cold treatment for the subsequent formation of flowers or for the hastening of flowering (6, 10). A plant in which flowering behavior is influenced by cold treatment applied in the seedling stage is said to be vernalizable.
McKee (5) has shown that a cold treatment during germination will hasten flowering in peas. Despite this evidence, peas have not generally been recognized as being naturally vernalizable. Peas have, however, been used extensively as experimental material in studies of the effect of various chemicals and growth substances on flowering. Leopold and Guernsey (7) found that the application to Alaska pea seeds of low concentrations of auxin (NAA) combined with a short cold treatment is effective in decreasing the number of nodes to the first flowers. Others, notably Fries (1) and Haupt (3, 4), have shown that the application to seeds or to seedlings of substances other than auxins can likewise affect the flowering of peas in terms of time to flower or in terms of the number of nodes to the first flower.
It will be shown in this paper that at least two varieties of peas, both late varieties, are naturally vernalizable, in the sense that their subsequent flowering behavior is influenced by a cold treatment given during the seedling stage. It will also be shown that the cold treatment has a distinct and separable effect on the vegetative development of the plant as well as on flowering, and that the pea is vernalizable only over a rather limited range in the physiological development of the plant. These facts make this plant an experimentally interesting one.
THE EFFECT OF A COLD TREATMENT (TWO EXPERIMENTS) ON THE
FIRST NODE TO FLOWER IN TWO VARIETIES OF PEA (PISUM SATIVUM L.)
|Days of Cold
|Varieties of Pea|
|5||18.4 ± 0.52||19.2 ± 0.46||17.0 ± 0.74 *||18.6 ± 0.86|
|10||17.3 ± 0.71 *||19.0 ± 0.75||16.5 ± 0.97 *||17.5 ± 0.92 *|
|20||16.2 ± 0.82 *||18.5 ± 0.53||15.0 ± 0.71 *||16.8 ± 0.45 *|
|30||16.0 ± 1.11 *||19.1 ± 0.9||14.5 ± 0.75 *||17.0 ± 0.75 *|
|0||18.63 ± 0.52 **||19.5 ± 1.0||18.2 ± 0.42 **||18.17 ± 0.85|
|5||17.7 ± 0.47 *||18.57 ± 0.79||18.4 ± 0.55||19.2 ± 0.73|
|15||16.8 ± 041 *||18.0 ± 0.82||16.57 ± 0.79 *||17.25 ± 0.5|
|25||16.0 ± 0.57 *||17.13 ± 0.53 *||153 ± 0.68 *||16.8 ± 0.45|
|Values given are the
means (± the standard error) of a minimum of 9 plants per treatment.
*Difference from controls significant at the 1% level.
** No significant difference between vernalized and devernalized controls.
The effect of a cold treatment on the vegetative development of plants, has been shown, for example, in Petkus winter rye (8) where vernalization results in decreased formation of tillers. However, it has not been shown whether the vegetative effect is independent of flowering or whether it is a secondary reaction dependent on the effect of cold treatment on flowering.
MATERIALS AND METHODS
Two pea varieties, Unica and Zelka (commercial varieties obtained from Holland) were used in these experiments. Both are late varieties which when grown under optimal greenhouse growing conditions of a long day (16 hrs) and relatively cool temperatures (17 to 20°C) will flower after the 18th node.
Seeds were soaked in tap water for from 4 to 6 hours at 20°C and planted in containers filled with a mixture of 50% vermiculite and 50% crushed rock. The containers were then put into constant temperature rooms at either 4 or 7°C in the dark for periods up to 30 days and watered when necessary with Hoagland's nutrient solution.
|FIG. 1. The effect of a cold treatment on the first node to flower. The broken curves show the effect of a high temperature treatment of 30°C for 10 days immediately following the cold treatment.||FIG. 2. The effect of a cold treatment on the height (in cm) to the first flowering node. The broken curves show the effect of a high-temperature treatment of 30°C for 10 days immediately following the cold treatment.|
Series of 30 plants were removed at intervals. In each series half of the plants were put directly into the normal growing conditions, which in these experiments consisted of 8 hours of natural light at 20°C, 8 hours of artificial light at 17°C and 8 hours of darkness at 17°C. The remaining plants of a series were maintained under devernalizing conditions of 30°C (16 hours of artificial light) for 10 days. At the end of the 10-day devernalizing treatment the plants were put into the normal growing conditions described above.
Measurements of flowering behavior were made by counting the number of nodes formed before the first flower appeared. Measurements of the vegetative development of the plants were made by measuring the height from the first bract to the first flowering node. Data were recorded after the internode below the flowering node had completely elongated.
The results of two of a series of experiments showing the effect of a cold treatment on the first node to flower are presented in table I and figure 1. Replication in each of these experiments was from 9 to 15 plants per treatment. Variability is expressed as the standard error of the mean. The two series shown are representative of the results obtained in other similar experiments.
For both varieties cold treatments of 5 days or more decreased the number of nodes to the first flower as compared with the control. The differences are statistically significant at the 1% level. It can also be seen from this table that the devernalizing treatment significantly delayed flowering as compared to the control. While in the variety Unica, the heat treatment which followed the vernalizing treatment completely reversed the effect of cold; in the variety Zelka complete reversal did not occur although the heat treatment following a cold treatment did significantly delay flowering.
|FIG. 3. The effect on the first node to flower of a pretreatment at 20 or 26°C prior to a cold treatment at 7°C for 25 days.|
Height to the first flowering node may be used as another measure of vernalization. That there is an inductive effect of a cold treatment on the height to the first flowering node can be seen from figure 2. In both varieties devernalization results in some increase in the height to the first flowering node but high temperature treatment following vernalization is apparently not as effective on vegetative development as it is on flowering. This is especially true when we consider the variety Unica in which the high temperature treatment completely reverses vernalization with respect to flowering.
|FIG. 4. The effect on the height to the first flowering node of a pretreatment at 20 or 26°C prior to a cold treatment at 7°C for 25 days.||FIG. 5. Average internode length as a function of a pretreatment at 20°C.|
In a second series of experiments plants were germinated at a high temperature prior to cold treatment in order to determine whether there was a critical time in the development of the pea seedling during which it was most receptive to a cold treatment. These plants were germinated at 20 or 26°C for periods up to 5 days. At the end of this time the seedlings were given a cold treatment at 4 or 7°C for 25 to 30 days; a period of cold sufficient to result in a maximum effect of the cold.
Figure 3 shows the results of one such experiment in which seeds of the variety Zelka were used. It is apparent that the longer the seeds are germinated at 20 or 26°C, the less effective is the following cold treatment in reducing the number of nodes to the first flower. With a 20°C pretreatment the critical period lies somewhere between 3 to 5 days since after three days at 20°C the plants can no longer be vernalized. When the plants are germinated at 26°C they remain vernalizable for 1 to 2 days. Three or more days of pretreatment at 26°C prevents subsequent cold treatment from exerting an effect on flowering.
The initial germinating temperature has no apparent effect on the height to the first flower. Figure 4 shows the results of a high temperature pretreatment on height to flower. These are the same plants from which the flowering data of figure 3 were taken. The decrease in height of the vernalized plants as compared to the unvernalized plants remains the same throughout the range of the pretreatment periods.
If the data of figure 4 are plotted as in figure 5, i.e., ratio of height to the first flowering node to the node at which the first flower appears, then it can be seen more clearly that flowering and vegetative development are independently affected by vernalization. If the decrease in height was due only to the decrease in the number of nodes to the first flower, the slopes of the hoes should be zero when actually they are less than zero. The figures on this graph can also be read as the average node length.
The results of these experiments show that peas, at least the varieties used in these experiments, can be vernalized and that the inductive effect of cold treatment can be reversed by high temperature treatment (i.e., they can be devernalized). It is interesting to note that in the 1948 report from the Netherlands of the Rijksinstituut voor Rassenonderzoek van Landbouwgewassen (9) on field tests of pea varieties, both varieties used in these experiments (Zelka and Unica) are considered to be relatively early varieties, flowering at the 12.9 and 14.5 nodes respectively. Since peas are normally planted early in spring when the temperatures are sufficiently low to allow the vernalization process to be consummated, it seems likely that vernalization has in fact actually occurred in the reported field trials.
In addition to the fact that the pea varieties can be vernalized and devernalized two other facts of interest have emerged:
1) There appears to be a distinct and separable effect of cold treatment on the vegetative development of peas, an effect which, like the effect of flowering, is inductive. Devernalization only partially reverses the effect of vernalization on vegetative development in the variety (Unica) in which devernalization completely reverses the vernalization with respect to flowering. In the variety Zelka this is not as clear, although the degree of reversal due to high temperature is far less for vegetative development than it is for flowering. Both the processes effected by a cold treatment, while obviously going on simultaneously must be distinct. This is especially apparent in the experiments in which the germinating seeds are pretreated at 20 or 26°C prior to receiving the vernalizing treatment. The fact that pretreatment of the germinating seed for a short period (3 to 5 days) prior to a cold treatment makes the plant insensitive to cold with respect to flowering, although it remains sensitive to a cold treatment with respect to the vegetative development, suggests that the effect of a cold treatment can be separated into a) a vegetative effect, and b) a reproductive effect, both of which are inductive. The former could be termed vegetative vernalization as opposed to vernalization which usually refers to the inductive effect of a cold treatment on flowering alone.
2) The high temperature pretreatment experiments indicate that there are physiological limits during which flowering can be effected by a cold treatment. With a pretreatment at 20°C of germinating seeds for a period up to 3 days, vernalization is still possible. With a pretreatment at 26°C for one day vernalization is still possible. But, with pretreatments of more than 3 days at 20°C and more than one day at 26°C vernalization is no longer possible. This is significant when one considers that dissection of growing points of samples of plants pretreated at 20°C up to 5 days, reveals no difference in the number of nodes from the number present in the dry seeds. At 26°C there is no change in the number of nodes present up to the fourth day; only elongation of the epicotyl occurs during these periods at both temperatures.
Since no new nodes have been laid down during the critical period of the pretreatments, some physiological changes rather than morphological changes must take place during this time. The plants appear to have passed through a critical stage in their physiological development such that they can no longer be vernalized.
These experiments seem to delimit the period in development which is most critical in studies of vernalization and indicate the developmental stages which should be used in studies of the biochemical changes occurring during the vernalization of peas.
The inductive effect of a cold treatment on flowering in peas has been studied. It has been found that the two late varieties studied are normally vernalizable with respect to both flower formation and vegetative development and that these two phenomena can be separated.
A pretreatment during germination at 20 or 26°C for up to 5 days prior to the optimal cold treatment results in a progressive loss of ability to be vernalized. Plants remain vernalizable for up to 3 days at 20°C. At 26°C they remain vernalizable for only 1 or 2 days.
The pretreatment at either temperature has no effect on vernalization with respect to vegetative development.