Leaf Length Tracker: a novel approach to analyse leaf elongation close to the thermal limit of growth in the field

Abstract

Leaf growth in monocot crops such as wheat and barley largely follows the daily temperature course, particularly under cold but humid springtime field conditions. Knowledge of the temperature response of leaf extension, particularly variations close to the thermal limit of growth, helps define physiological growth constraints and breeding-related genotypic differences among cultivars. Here, we present a novel method, called 'Leaf Length Tracker' (LLT), suitable for measuring leaf elongation rates (LERs) of cereals and other grasses with high precision and high temporal resolution under field conditions. The method is based on image sequence analysis, using a marker tracking approach to calculate LERs. We applied the LLT to several varieties of winter wheat (Triticum aestivum), summer barley (Hordeum vulgare), and ryegrass (Lolium perenne), grown in the field and in growth cabinets under controlled conditions. LLT is easy to use and we demonstrate its reliability and precision under changing weather conditions that include temperature, wind, and rain. We found that leaf growth stopped at a base temperature of 0°C for all studied species and we detected significant genotype-specific differences in LER with rising temperature. The data obtained were statistically robust and were reproducible in the tested environments. Using LLT, we were able to detect subtle differences (sub-millimeter) in leaf growth patterns. This method will allow the collection of leaf growth data in a wide range of future field experiments on different graminoid species or varieties under varying environmental or treatment conditions.

Keywords: Field conditions; leaf elongation; low temperature; marker tracking; monocotyledons; phenotyping; plant growth..

Fig. 1.

Experimental set-up of the measurement panel in the field. Similar panels without a roof were used in climate cambers. (A) Row-wise measurement of 20 wheat leaf replicates. Here, it is visible how threads are passing the first and second reverse rollers. (B) Close up view of a leaf tip attached by a hairpin. (C) The 20g counterweight and third reverse roller. (D) Wheat micro plots and the installed measurement panels with white beads and a near-infrared camera in front. The first three beads on the left were used for reference measurements. The panel was north facing to avoid shading of the investigated plants. The panel roof was installed to prevent confounding effects by raindrops and snow on the beads.

Fig. 2.

Linear correlation of image based displacement measurement against the manual calliper controlled movement of a bead. Similar correlations were found at each of the 23 panel positions.

Fig. 3.

Time series of week 1 of leaf length measurements in four wheat varieties from 26th March to 2nd April 2014. Measurements were taken from leaf 7 when it first emerged until the leaves were fully developed. The upper graph is the mean cumulative leaf length of n = 20 leaves per wheat variety. The middle graph shows the corresponding mean LER. The lower graph shows the corresponding air temperature measured at 5cm above ground and the soil temperature measured 5cm below ground. Grey stripes indicate night hours.

Fig. 4.

Linear correlations of mean LER (each n = 20 leaves) and air temperature at 5cm above ground for each of the four wheat varieties. Grey dots refer to week 1 (leaf 7) and black triangles to week 2 (leaf 8) of the experiment. The histograms in the upper left corners show LER corrected for thermal time (LER − aT) for week 1 and 2. Note: there is no data for week 2 for variety ‘Combin’.

Fig. 5.

Upper row: LER per °C (a) of three summer barley varieties (each n = 7 leaves) from the two field sites and the climate chamber. P-values are derived from ANOVA and letters above boxes indicate significant genotype-specific differences (Tukey-Kramer HSD, P < 0.05). Lower row: Histograms of mean temperature per hour (in steps of 0.5°C) from the period of measurement.