J.PlantPhysiol. Vol. 138.pp. 661-666{1991}

Introduction

The Effect of Low Root Temperature on Growth and Lipid Composition of Low Temperature Tolerant Rootstock Genotypes for Cucumber

H. A. M. BULDER, A. P. M. DEN NIJS!, E. J. SPEEK, P. R. VAN HASSELT, and P. J. C. KUIPER

University of Groningen, Department of Plant Physiology, P.O. Box 14,9750 AA Haren, The Netherlands

1 Centre for Plant Breeding Research, P.O. Box 16,6700 AA, Wageningen, the Netherlands

Received January 31,1991 . Accepted May 18, 1991

Summary

In the framework of research directed to diminish energy consumption of glasshouse cucumber pro­duction, three low temperature tolerant rootstock genotypes for cucumber were compared. Firstly, growth at low root temperature of one Cucurbita ficifolia and two Sicyos angulatus genotypes was studied to determine which was the one most suitable as a cucumber rootstock at suboptimal root tem­peratures. Secondly, differences in lipid composition were examined. Thirdly, lipid composition of these rootstock genotypes was compared with that of low temperature sensitive cucumbers (Cucumis sativus 1.; cv. Farbio and CPO inbred lines 79345 and 81354) studied earlier, to determine whether observed dif­ferences in lipid composition were consistent with differences in growth at low temperature of related species.

Plants were grown at an air temperature of 20°C d/12 °C n and at constant root temperatures of 20°C, 16 °C or 12°C. Although growth decreased at 12 °C for all genotypes, both Sicyos angulatus genotypes were more tolerant to low root temperature than Cucurbita ficifolia. Low root temperature affected root lipid composition only. An increased phospholipid and a markedly lower sterol and sterol ester level re­sulted in a strongly decreased sterol/phospholipid ratio at 12°C. Growth at low root temperature was in­versely related with the sterol/phospholipid ratio but no correlation between growth and sterol/ phos­pholipid ratio of the rootstock genotypes within one temperature regime was observed. So, this ratio is not an appropriate selection criterion for growth capacity.

Ke')I wor.ds: Cucurbita ficifolia, Cucumis sativus, fatty acids, phospholipids, phosphatidyl glycerol, root tem· perature, Sicyos angulatus, sterols.

Abbreviations: C. ficifolia (Cf) = Cucurbita ficifolia; CPO = Centre for Plant Breeding Research; LAR = leaf area ratio; NAR = net assimilation rate; PG = phosphatidyl glycerol; RGR = relative growth rate; S. angulatus (Sa) = Sicyos angulatus.

Low temperature and low light intensity acclimation are of great importance in cultivating glasshouse vegetable crops, especially in winter when much energy is needed for the pro­duction of crops like cucumber, tomato and sweet pepper. Since 1974, The Centre for Plant Breeding Research (CPO,

formerly the Institute for Horticultural Plant Breeding (IVT), Wageningen, the Netherlands) has been screening for less energy consuming cucumber and tomato genotypes (den Nijs, 1980 a; Smeets and Garretsen, 1986; den Nijs and Smeets, 1987).

Two of the resulting cucumber genotypes and a control va­riety, differing in genetic adaptation to low energy condi-

© 199! by Gustav Fischer Verlag, Stuttgart

662 H. A. M. BULDER, A. P. M. DEN NIJS, E. J. SPEEK, P. R. VAN HASSELT, and P. J. C. KUIPER

tions (Bulder et al., 1987), have also been screened for lipid differences, because membrane lipid composition is a com­monly used parameter for cold tolerance (Smolenska and Kuiper, 1977; Horvath et al., 1980; Vigh et al., 1985). How­ever, hardly any genotypic differences in lipid composition were observed in the two cucumber genotypes mentioned (Bulder et al., 1990), even though several examples of lipid changes as a result of low temperature, or differences in lipid composition of plants with different cold or chilling resist­ance, have been described in the literature (Lyons and Brei­denbach, 1979). The genetic differences might have been too small, or the genotypes may have lacked the capacity of lipid adaptation to low temperature. Differences in growth re­sponse to low temperature were evident for instance in the shoot/root ratio, in which respect plants that performed well under low energy conditions showed a high value (Bulder et al., 1987), indicating that low temperature tolerant genotypes had a more efficient root system at low tempera­ture than sensitive ones.

In outdoor-cucumber culture practice and also in late spring glasshouse cultivation, cucumbers (Cucumis sativus L.) are often grafted on the rootstock of Cucurbita fici/olia (figleaf gourd) because of its resistance to soil pathogens. The growing season of grafted cucumbers is also lengthened (Vf­felen, 1984). In Japan, the rootstock Sicyos angulatus (bur cu­cumber) is used for this purpose in glasshouse cucumber cul­tivation on cold soils (Tachibani, 1982).

Den Nijs (1980b, 1984a, b) showed improved growth of grafted plants of several cucumber genotypes on C. fici/olia and S. angulatus at low root temperatures. After grafting on C. fici/olia, Tachibani (1982) also found an improved per­formance of a Japanese summer cucumber variety exposed to low root temperature. The use of a low temperature tol­erant rootstock seems promising in saving energy in winter glasshouse cultivation. Several cucurbit species and geno­types from all over the world were collected to test their ca­pacity to serve as a rootstock for improved growth at low temperature (Visser and den Nijs, 1987).

In the present investigation growth of Cucurbita fici/olia, together with two genotypes of the bur cucumber, Sicyos angulatus, was studied at normal and low root temperature. We were also interested whether differences in growth would be reflected in lipid composition. Furthermore, we compared the lipid composition of the rootstock genotypes with the lipid composition of the glasshouse cucumber geno­types studied earlier (Bulder et al., 1990). The comparison concerned low temperature sensitive cucumber, low temper­ature tolerant cucumber and the low temperature tolerant rootstock genotypes. Analysis of leaf phosphatidyl glycerol (PG) was included since PG has often been mentioned as a selection criterion for chilling tolerance (Murata, 1983).

Materials and methods

Plant material

The plant material consisted of two Sicyos angulatus (Sa) geno­types and the commercially available Cucurbita ficifolia (C£). The small seeded S. angulatus (Sa I) was obtained from Royal Sluis Seed Company as KJ-100. The large seeded one (SA II) was acquired from

the Botanical Garden in Toronto and is maintained at the CPO as P 83007. It is part of a collection of over 20 accessions of Sicyos as­sembled for further evaluation as a rootstock for cucumbers under glass (Visser and den Nijs, 1987).

Growth experiment

Analysis of growth was carried out in a hydroponic system in a climate room at the CPO, Wageningen. The hard seed coat (actually the fruit wall) of the Sicyos seeds was partly removed and seeds were incubated in a refrigerator at 4°C during 6 days in order to improve and synchronize seed germination (Visser and den Nijs, 1987). Seed weight of the three genotypes was determined and seeds were plant­ed in trays with vermiculite saturated with standard Steiner's nutri­ent solution at 22°C constant (Steiner, 1968). After 7 days, the seed­lings were washed free of vermiculite and distributed over double­walled insulated tanks (200 x 40 x 40 cm). Per temperature regime, four tanks were connected by an overflow to a central storage con­tainer with Steiner nutrient solution in which the temperature was automatically adjusted. The solution of one group of tanks was kept at 20°C and that of the other group at 12°C by a a system of forced heating and cooling. A third group was kept at a root temperature of 16°C. Circulation of the nutrient solution was provided by pumps, and a 30 cm free fall of the nutrient solution through the overflow system assured sufficient aeration.

The plants in the tanks were exposed to about 22 W m - 2 visible radiation (SON-T lamps) for 8 hours daily. Air temperature was maintained at 20°C day/12°C night.

Experimental set·up

In a single tank, plants of each genotype were introduced in one field plot of 3 and one of 4 plants. One plant of each plot was harvested at 3 consecutive dates, while the fourth plant was used for lipid analysis. In this way, 8 replicate field plots for growth were ob­tained for each genotype at each root temperature treatment. Har­vest took place 30, 38 and 46 days after sowing. Stem length, num­ber and total area of the leaves and fresh and dry weights of both shoots and roots for each individual plant were determined. For the leaf area measurements aLI-COR model 3100 area meter was used.

The growth data were analysed at CPO using a MANOV A pro­cedure that was especially developed for discriminating genotypes with regard to growth parameters (Keuls and Garretsen, 1982; Gar­retsen and Keuls, 1986). In the present contribution only the most relevant data of the 20 and 12°C root temperature treatment are presented for comparison with the lipid results.

Lipids

For lipid analysis 4 replicates were obtained around the last har­vest date of the growth experiments for 12 °C and 20°C root tem­peratures. Lipids were extracted and lipid composition was de­termined as described earlier for cucumber (Bulder et al., 1990).

Results

Growth Analysis

The results were transformed in a MANOV A procedure. For illustrating the genotype differences means of the curves obtained are most suitable.

At 20°C root temperature, total mean dry weight of all three genotypes differed significantly, Cf having the highest and Sa I having the lowest dry weight. Low root temperature

drastically reduced dry weight of all genotypes (Table 1). At 12°C root temperature, Cf and Sa II had significantly higher dry weights than Sa I.

For RGR there were no differences between the genotypes at the 20°C temperature regime. RGR was reduced by 12°C root temperature for Cf and Sa I, but not for Sa II. Both Sa genotypes maintained a higher RGR than Cf. For NAR the only significant genotypic difference was observed at 20°C root temperature, where Sa I had a higher NAR than Cf. Under the 20°C regime, Cf had a higher LAR than the two Sa genotypes. At 12°C root temperature, LAR of all geno­types was lower than at 20°C and most drastically so for Cf; both Sa genotypes had a higher LAR than Cf.

Lipids

Leaves (Table 2)

Neither at 20 nor at 12°C root temperature, were genetic differences in total lipid/ g fresh weight observed. The contri­bution of phospholipids to total lipids varied with the geno­type. At 20°C root temperature, the phospholipid/total lipid ratio was significantly different between the two Sa genotypes, and at 12 °C root temperature between Cf and Sa II. For both temperature regimes Sa II had the highest phos­pholipid content. Under both root temperature regimes, total fatty acid level was significantly higher for Sa II as com­pared with Cf and Sa I. At 20°C root temperature, the de­gree of unsaturation was lower for Sa I than for Sa II and Cf. At 12°C no genotypic differences were observed.

The effect of low root temperature was manifest in the phospholipid to total lipid: Sa II had less phospholipids, and the fatty acids were more saturated under these conditions. Sa I had a high and Sa II had a low degree of unsaturation of fatty acids.

Table 1: Growth characteristics of C. ficifolia (Cf) and 2 S. angulatus (Sa I and Sa II) genotypes under optimal (20°C) and suboptimal (12°C) root temperature regimes. Values are mean levels of the curves derived from the MANOVA's. Values in the same column, followed by the same letter, are not significantly different at the 5 % level.

Genotype 20°C 12°C

Total dry weight, g Cf 1.57 a 0.90 a Sa I 0.72 c 0.45 b Sa II 1.13 b 0.87 a

RGR, g g-I day-I Cf 0.123 a 0.080 a Sa I 0.142 a 0.105 b Sa II 0.120 a 0.125 b

NAR, mg (cm2)-1 day-I Cf 0.276 a 0.317 a Sa I 0.372 b 0.338 a Sa II 0.308 ab 0.393 a

LAR, cm2 (gt 1

Cf 446 a 261 a Sa I 381 b 314 b Sa II 386 b 319 b

Low root temperature, growth and lipid composition 663

Table 2: The effect of optimal (20°C) and suboptimal (12 0C) root temperature regimes on different leaf and root membrane lipid frac­tions of C. ficifolia (Cf) and S. angulatus I and II (Sa I and Sa II). Values in the same column, followed by the same letter, are not sig­nificantly different at the 5 % level. Comparison of optimal and sub­optimal root temperature: "*, ", significant at 1 %, 5 % respectively; n.s., not significant; n.d., not determined (Student t-test n = 4).

Genotype Leaf

20°C 12°C Total lipid, mg (g fresh weight)-I Cf 11.9 a 15.8 a" Sa I 15.0 a 17.7 a n.s. SaIl 11.9 a 14.2an.s.

Phospholipid, nmol P (mg lipid)-I Cf 274 ab 245 a n.s. Sa I 244 a 251 ab n.s. Sa II 315 b 270 b*

Fatty acids, /lg (mg lipid) - 1

Cf 363 a Sa I Sa II

333 a 411 b

387 a n.s. 383 a" 435 b n.s.

Degree of unsaturation of fatty acids Cf 2.38 a 2.36 a n.s. Sa I 2.31 b 2.35 a* Sa II 2.38 a 2.33 a*"

Free sterols + sterol esters, /lg (mg lipid)-I Cf n.d. n.d. Sa I n.d. n.d. Sa II n.d. n.d.

Roots (Table 2)

20°C

5.2 a 2.9 b 3.1 b

490 a 665 b 685 b

260 a 381 b 392 b

1.78 a 1.74 a 1.78 a

146 a 182 b 180 b

Root

12°C

6.2 a" 4.4 b" 4.3 b*

690 a"" 835 b" 821 b"

383 a*" 510 b*" 500 b*"

1.96 a"* 1.94 a** 1.97 a"*

126 a* 114 a"" 123 a*"

At both 20 and 12°C root temperatures, Cf significantly differed from both Sa genotypes. Cf had more total lipid, less phospholipids and less fatty acids than the Sa genotypes. The degree of unsaturation of the fatty acids showed no geno­typical differences under either root temperature regime. The sterol contents only differed at 20°C root temperature, Cf having less sterols.

As a result of low root temperature, total lipid/ g fresh weight, phospholipidltotal lipid, fatty acidltotal lipid and the degree of unsaturation of fatty acids were increased. At the same time, the sterolsltotal lipid ratio was strongly de­creased. The increase in phospholipids, induced by low tem­perature, and the decrease in free sterols + sterol esters re­sulted in a strong decrease in sterol/phospholipid ratio at 12°C root temperature (Table 4, lower part).

Effect of low root temperature on leaf phosphatidyl glycerol

Genotypic differences were observed under both root tem­peratures (Table 3). Cf had less PG per total lipid than Sa II. The fatty acid composition of PG differed at 20°C root tem­perature for all genotypes; Cf had the highest % of relatively saturated fatty acids (16: 0 + trans, 3 -16: 1, Murata, 1983; 16: 0 + trans, 3 -16: 1 + 18: 0, Bishop, 1986). At 12 DC, Cf and Sa I differed from Sa II, which had the lowest % of rela­tively saturated fatty acids. The degree of unsaturation of leaf PG was different for all genotypes at both root temperatures

664 H. A. M. BULDER, A. P. M. DEN NIlS, E.]. SPEEK, P. R. VAN HASSELT, and P. J. C. KUIPER

Table 3: The effect of optimal (20°C) and suboptimal (12 0C) root temperature on the amount of leaf PG and the composition and un­saturation of its fatty acids in C. ficifolia (Cf) and S. angulatus I and II (Sa I and Sa II). Values in the same column, followed by the same letter, are not significantly different at the 5 % level. Comparison of optimal and suboptimal root temperature: *, significant 5 %; n.s., not significant (student t-test n = 3 for C. ficifolia and n = 4 for both S. angulatus genotypes).

----------------------------Genotype 20°C 12 °C

PG fatty acid, J'g (mg lipid)-I Cf 36.0 a 40.2 a* Sa I 41.7 ab 42.8 ab n.s. Sa II 45.4 b 45.5 b n.s.

(16:0 + trans, 3-16: 1) PG, % Cf 57.7 a 56.2 a n.s. Sa I 50.7 b 54.0 a n.s. Sa II 49.1 b 49.4 b n.s.

(16 :0+trans, 3-16:1+18:0) PG, % Cf 61.2 a 60.4 a n.s. Sa I 52.4 b 56.0 a n.s. Sa II 50.8 b 51.1 b n.s.

Degree of unsaturation of PG fatty acids Cf 1.33 a 1.32 a n.s. Sa I 1.46 b 1.44 b n .s. Sa II 1.59 c 1.56 c n.s.

and increased in the order Cf < Sa I < Sa II. Only for Cf, total PG was somewhat higher at 12°C than at 20 0c.

Discussion

Growth

Both Cf and the Sa genotypes reacted to root temperature with differences in growth. At 20°C, Cf had a higher total dry weight than either of the Sa genotypes and this was not due to a higher RGR during the experiment (Table 1). The differences must reflect the size of the plants at the start of the experiment (Visser and den Nijs, 1987). Cf possessed a higher LAR than the Sa genotypes but this advantage was an­nihilated by a lower NAR. Similar mutual compensation ef­fects of NAR and LAR were earlier observed in tomato (Smeets and Garretsen, 1986) and in cucumber (Bulder et al., 1987; Den Nijs and Smeets, 1987).

Total dry weights of Cf and Sa II were alike at 12°C root temperature, while Sa I had a lower dry weight. For Cf and Sa I, the RGR was decreased by low root temperature but Sa II showed no reaction in RGR. The decrease in RGR was primarily due to a decreased LAR and not to a strongly de­creased NAR. There was hardly any root temperature effect on NAR in this range, and at 12°C root temperature no genotypic differences were observed. NAR seems to be rath­er stable for related species within the temperature range of 12 - 20°C (Den Nijs, 1980 a; Kleinendorst and Veen, 1983; Bulder et aI., 1987; Den Nijs and Smeets, 1987). The LAR at 20°C root temperature, was higher for Cf than for the Sa genotypes, but at 12°C it was strongly reduced for Cf, even below the LAR of the Sa genotypes. Obviously growth of Sa was less sensitive to low root temperature than that of Cf,

suggesting that especially at low root temperatures, Sa could be a promising rootstock.

Lea/lipids

The differences in growth between Cf and both Sa geno­types were hardly reflected in the leaf lipid composition of plants grown at 20°C and 12 °C root temperature. Neither was an effect of low root temperature on leaf lipids present, except for the degree of unsaturation of the leaf fatty acids, which was decreased in Sa II and increased in Sa I. These genotypes were least affected in growth by 12°C root tem­perature. The fatty acid results contradict those of Tachibani (1986), who found little influence of root temperature on fatty acid composition of the leaf lipids of two cucumber varieties and Cf.

To check whether the root temperature might have a specific effect on leaf lipid composition, the amount of PG and its fatty acid composition was also examined (Table 3). A distinction between low temperature sensitive Cf versus both Sa genotypes was clear, especially at 20°C root temper­ature. PG in leaves of Cf was more saturated (expressed as a higher 16: 0 + trans, 3 -16: 1 content (Murata, 1983) and 16:0 + trans, 3-16: 1 + 18:0 content (Bishop, 1986) and a lower degree of unsaturation), but the low root temperature effect on amount of PG and fatty acid unsaturation was ab­sent in all genotypes (except Cf: higher amount of PG). Ex­posure of the total plant of the cucumber variety Farbio to suboptimal temperature resulted in a decreased saturation of PG in the leaf lipid, whereas the relatively saturated fraction in PG decreased (Bulder et aI., 1990). The more low tempera­ture tolerant inbred lines of cucumber did not show such a temperature dependent change in PG. It, can be concluded that PG, which is associated with the chloroplast membrane, was not influenced by low root temperature. Photosynthesis of lucerne was not affected by changes in root or leaf temper­ature over a range of 5-25 °C (Harding and Sheehy, 1980); the relatively stable NAR values observed in both cucumber and the rootstock genotypes are in agreement with this ob­servation.

Root lipids

In general the genotypic differences in growth between Cf and the Sa genotypes could also be discerned in the lipid characteristics. Genotypical differences for both root tem­perature treatments were visible as higher total lipid/ g fresh weight, less phospholipidltotallipid and less fatty acid/total lipid in Cf roots than in Sa roots (Table 2). No genotypical differences in the degree of unsaturation of root fatty acids were observed, indicating that this root parameter was not related to shoot growth. Low temperature treatment re­sulted in an increased totallipidl g fresh weight, an increased phospholipid per g fresh weight and per total lipid, increased fatty acid/totallipid, a higher degree of unsaturation of the fatty acids and a decreased sterol/totallipid ratio. This result is in accordance with Tachibani (1986, 1987), who found an increase in total lipid and phospholipid/g fresh weight in cu­cumber and in Cf roots as the result of low temperature; and also with observations by Horvath et a1. (1983), who found

an increase in phospholipid/ g fresh weight in leaves of differ­ent cucumber genotypes at low growth temperature. The ef­fect of low temperature on the contribution of phospho­lipids to total lipids, found in the present investigation for all rootstock genotypes, was not present in cucumber (Bulder et al., 1990), where the ratio remained constant. Thus, related species obviously do not react in the same way to low root temperature, or the severity of the temperature stress differ­ed too much. The degree of unsaturation of the fatty acids in root lipids increased in both the rootstock genotypes and cu­cumber as a result of low temperature, as generally reported (Lyons and Breidenbach, 1979). Often, low temperature tol­erant species show a larger increase in fatty acid unsaturation than the sensitive ones (e.g. for broccoli and soybean, Markhart et al., 1980). In our investigations the rootstock genotypes also showed a larger increase in unsaturation than the more low temperature sensitive cucumber genotypes (this study, Bulder et al., 1990). The root sterol content of the rootstock genotypes was decreased by low temperature. The same effect was also reported for cucumber leaves by Horvath et al. (1983) but not for cucumber roots (Bulder et al., 1990).

A strong decrease in the root sterol/ phospholipid ratio was observed for all genotypes as a result of the strong in­crease in phospholipids and the decrease in sterols at 12°C root temperature (Table 4). Horvath et al. (1987) found a strong inverse relation between sterol/phospholipid ratio and frost resistance in leaves of cereals: the higher the ratio, the more frost sensitive the plasma membrane seemed to be. At a sterol/phospholipid ratio of 0.39, strongly expressed sterol-sterol interactions in lipid bilayers were observed in x­ray studies, indicating disruptions in the membrane whereas

Table 4: Sterol/phospholipid ratio's (on a molar basis) of roots of different cucurbits (cucumber and rootstock genotypes for cucum­ber), differing in genetic adaptation to low temperature and low light intensity, grown at different temperature regimes (optimal: 25 °C/20 °C daylnight for cucumber, 20 °C/12 °C daylnight and root 20°C constant for the rootstock genotypes and suboptimal: 20 °C/12 °C daylnight for cucumber and 20 °C/12 °C daylnight and root 12°C constant for the rootstock genotypes). (Cucumber data derived from Bulder et al., 1990) 1 mol sterol: 408.757 g, based on 14 % cholesterol; 9 % campesterol; 38.5 % stigmasterol and 38.5 % sitosterol (Bulder unpublished results) Cucumber genotypes: cv. Farbio, CPO line 79345 and CPO line 81354. Rootstock geno­types: C. fici/olia (Cf), S. angulatus I (Sa I) and S. angulatus II (Sa II).

Genotype Root

Optimal Suboptimal Sterol/phospholipid ratio

Cucumber shoot & root 25/20 DC, din 20/12 DC, din Farbio 0.770 0.762 CPO 79345 0.988 0.818 CPO 81354 0.926 1.071

Rootstock genotypes shoot 20/12 DC, din 20/12 DC, din

root 20°C, d & n 12°C, d & n Cf 0.729 0.446 Sa I 0.669 0.334 Sa II 0.642 0.367

Low root temperature, growth and lipid composition 665

at a sterol/phospholipid ratio of 0.08 no sterol-sterol interac­tions were observed, even at -10°C. When subjected to low temperature treatment, the roots of cucumber genotypes (Bulder et al., 1990) had a higher sterol/phospholipid ratio (0.762 -1.071) than the presently studied rootstock geno­types (0.334-0.729) (Table 4). When the present results are compared with the above mentioned cucumber data, it is evi­dent that the sterol/phospholipid ratio of root lipids de­creased in the following order: cucumber 25 °e/20 DC, cu­cumber 20 °C/12 DC, rootstock genotypes 20°C root temperature and rootstock genotypes 12 °C root tempera­ture. These results for roots correspond with the above men­tioned inverse relationship of Horvath et al. (1987) for frost resistance in leaves of cereals. It should be noted that the comparison holds only for root sterol/phospholipid ratio's. The leaf sterol/phospholipid ratio's of the cucumber geno­types observed by Horvath et al. (1983, 0.11- 0.17) do not fit in the above mentioned range, probably because the com­position of leaf lipids differs too much from that of root lipids. Within a single temperature treatment, the individual genotypes did not show a clear relation between growth re­sponse and sterol/phospholipid ratio.

In conclusion, there were genotypic differences in growth at low root temperature between Cf and both Sa genotypes, Sa being the rootstock genotype least sensitive to low tem­perature. The differences in growth were reflected in the root lipid composition. The sterol/phospholipid ratio show­ed a clear and consistent reaction to low temperature treat­ment, both in low temperature tolerant cucumbers and in the rootstock genotypes. However, no correlation between growth response and sterol/ phospholipid ratio was found with the rootstock genotypes used in this study, suggesting that the sterol/ phospholipid ratio is primarily an indicator of low temperature tolerance as such and not a suitable selec­tion criterion for growth potential of closely related geno­types at suboptimal temperature.

Acknowledgements

This investigation was a cooperation of the CPO, Wageningen and the RUG, Groningen. It was supported by the committee for Energy Consumption of the Dutch Ministry of Agriculture and Fisheries. We thank Dick Visser for carrying out the growth experi­ments, Dr. L. Smeets for carefully reading the manuscript and the statistical section of the CPO for the MANOV A's on the growth experiments.

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