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The chilling-sensitive genotypes were characterized by lower MDA and antioxidant contents than the chilling-tolerant genotypes, which suggests a role for oxidative stress tolerance as a part of chilling tolerance in miscanthus. Although numerous studies have linked chilling tolerance in maize to oxidative stress Fryer et al. One exception is the study by Ezaki et al. Here we have presented results for several indicators of redox homeostasis for a common set of five miscanthus genotypes.

In general, the lower chlorophyll content at the beginning of the growing season indicates that MDA and antioxidant contents were high in the early growing season relative to the chlorophyll content for all genotypes. It is therefore possible that, relative to the amount of light energy captured, more oxidative stress occurred at T1.

This observation agrees with the results of Fryer et al. Antioxidant defences might thus be important to chilling stress tolerance in miscanthus, but no disruption of the redox homeostasis was observed in our study. MDA levels did not change significantly over time in any of the genotypes. There was therefore no strong indication of higher lipid peroxidation on T1. Catalase activity and glutathione concentration were significantly higher in the beginning of the growing season, indicating an increased need for protection against reactive oxygen species ROS at T1.

The latter agrees with the observation of higher levels of antioxidants in chilling-tolerant maize genotypes than in sensitive genotypes Leipner et al. The amount and composition of carbohydrates in leaf tissues varied significantly among genotypes and sampling dates.

The chilling-sensitive genotypes OPM35 and OPM51 were similar in carbohydrate composition and were characterized by high levels of glucose and fructose as well as a high glucose to sucrose ratio. According to available literature, maltose and raffinose are the sugars induced most in plants under chilling stress Tarkowski and Van den Ende, and have been shown to act as protective agents for cell membranes Kaplan and Guy, ; Valluru and Van den Ende, , act as antioxidants Nishizawa et al. In agreement with this view, maltose concentrations were inversely related to temperature, with a clear tendency to decrease throughout the study period in all genotypes except OPM66, for which maltose concentrations were not the highest at T1.

These higher concentrations of maltose on T1 could be a protective measure against chilling stress. On the other hand, while raffinose concentrations were the lowest in all genotypes at T3, the warmest sampling point, they increased again at T4 and T5, resulting in no clear overall relationship between raffinose concentration and temperature. This contrasts with the findings of Fonteyne et al. Overall, the chilling-tolerant OPM06 and OPM09 had higher concentrations of raffinose than other genotypes throughout the sampling period. Souza et al. This agrees with knowledge available in rice Oryza sativa , oat Avena sativa and Arabidopsis thaliana , where raffinose contents are higher in chilling-tolerant genotypes Klotke et al.

However, raffinose concentrations were not significantly higher in the chilling-tolerant genotype OPM66 at colder sampling dates, indicating other responses in this genotype. Taken together with the results for maltose concentration, these results indicate that, in regard to sugars, the protection mechanisms and metabolic responses of genotype OPM66 is different from those of other relatively chilling-tolerant genotypes such as OPM06 and OPM Analysis of the overall response of genotypes and temporal patterns using PCA indicated that sampling date was the main differentiating factor, with T1 clearly separated from warmer sampling moments.

As anticipated by the results obtained for the different parameters separately, OPM09 took an intermediate position in the PCA plot. We can therefore conclude that the genotypes chosen as chilling sensitive and the genotypes chosen as chilling tolerant differ at the metabolic level, even if they might differ for particular types of reactions as illustrated by differing trends for specific biochemical characteristics. Of particular interest are the results obtained for OPM while in the beginning of the growing season this genotype was most similar to the chilling-tolerant genotypes, after the warm weather around 12 May , it was more similar to the chilling-sensitive genotypes, with higher fructose and glucose contents and higher PPDK activity.

Of all investigated traits, WSC analysis was the most informative, as it indicates both chilling tolerance and growth capacity. The analysis of WSCs was also fast and relatively easy to perform.

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Purdy et al. Sampling at midsummer, as done by Purdy et al. The ratio of glucose to sucrose clearly distinguished the chilling-tolerant and chilling-sensitive genotypes in our study; if this would be confirmed, this ratio could be a good marker trait to select chilling-tolerant genotypes. The overall physiological response of chilling-tolerant genotypes was clearly distinguishable from that of chilling-sensitive genotypes.

Chilling-tolerant genotypes accumulated protective monosaccharides such as raffinose and sucrose and displayed high catalase activity at low temperatures.

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The chilling- sensitive genotypes were characterized by higher concentrations of glucose and fructose, and higher PPDK activity later in the growing season, indicating a higher photosynthetic activity. Overall, there appeared to be a trade-off between high growth and chilling stress tolerance for the investigated genotypes.

Of all the traits measured, WSCs appear to be the most suitable for large-scale screening, since these were both fast to measure and had a strong relationship to chilling tolerance.

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It thus appears to be possible to combine both chilling tolerance and strong growth in one genotype. We also want to acknowledge Wageningen University and Aberystwyth University for making available the miscanthus genotypes. Aebi H. Catalase in vitro.

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