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Transpiration efficiency in a segregating population of sunflower: Inheritance, correlation with other traits and association with hybrid grain yield

Christopher J. Lambrides1, Scott Chapman2 and Ray Shorter3

1 University of Queensland School of Land and Food Sciences, St Lucia 4072, Brisbane, Australia.
Email chris.lambrides@uq.edu.au

2,3 CSIRO Plant Industry, Queensland Bioscience Precinct, 306 Carmody Rd, St Lucia 4067, Brisbane, Australia

Abstract

Plants incorporate isotopes of carbon into their tissue at different rates because of discrimination against 13C relative to 12C during photosynthesis. This difference in discrimination has been negatively correlated with transpiration efficiency (TE) in many C3 species and so, carbon isotope discrimination (Δ) of leaf tissues has been proposed as a potential tool for selecting genotypes with improved performance under water limited conditions. The relationship between Δ and TE in sunflower has been described previously using diverse genotypes, but this relationship has not been investigated with material selected from a segregating population. In this study, the TE of twenty recombinant inbred lines from a population (HAR4 x SA52) segregating for Δ was evaluated in a rainout shelter experiment. A strong negative genetic correlation between TE and Δ was observed (rg = -0.58), confirming previous studies of sunflower with unrelated lines. In addition, TE was strongly correlated to plant height at the final harvest (rg = 0.64) and TDW (rg = 0.58), and moderately correlated to SLW (rg = 0.46) and SPAD (rg = 0.21) but not leaf number (rg = 0.02). Estimates of narrow sense heritability of TE and Δ were very high (0.82 and 0.77, respectively) suggesting that selection for these traits could occur in early generations of segregating populations. Grain yield evaluations under field conditions of hybrids contrasting for Δ showed that low Δ (high TE) hybrids had a yield advantage between 22-35% in dry environments where the yield was less than 2t/ha. While this level of yield advantage may not be realized in commercial breeding programs, computer simulations suggest that 10-15% yield improvements may be possible. Low Δ material selected from the population HAR4 x SA52 has been distributed to private seed companies for further evaluation.

Media summary

Sunflower germplasm with higher transpiration efficiency (TE) has been identified. Carbon isotope discrimination is a useful selection tool for TE in sunflower breeding programs.

Keywords

Carbon-isotope-discrimination, isotopes, Helianthus annuus, abiotic stress, drought tolerance

Introduction

In Australia, the majority of sunflower is grown in the summer in dry subtropical environments (<500 mm rainfall) on soils that vary in plant available water holding capacity from 90 to 250 mm. These production environments are also characterised by unreliable rainfall across years, large evaporative demands and maximum temperatures frequently exceeding 35C. Consequently average yields in Australia are relatively low at approximately 990 kg/ha (1993-2002; FAOSTAT 2003). Water is the major abiotic limitation to growth and yield and its efficient use by dryland crops is an important adaptation attribute.

Plants incorporate isotopes of carbon into their tissue at different rates because of discrimination against 13C relative to 12C. This difference has been negatively correlated with transpiration efficiency (TE – the amount of dry matter produced per unit transpiration) in many C3 species and so carbon isotope discrimination (Δ) of leaf tissues has been proposed as a potential tool for selecting genotypes with improved performance under water limited conditions (Farquhar and Richards 1984; Hubick et al. 1988; Condon and Richards 1992; Condon et al. 1993; Rebetzke et al. 2002). Rebetzke et al. (2002) demonstrated that early generation divergent selection for Δ affected grain yield among high and low Δ, BC2F4:6 progeny of wheat (Triticum aestivum L.). The low Δ lines had a yield advantage compared to the high Δ lines of up to 11% in low rainfall Australian environments.

Generally, a negative correlation exists between leaf Δ and TE (see review by Hall et al. 1995). Virgona and Farquhar (1996) observed a negative correlation (r = -0.98, P<0.001) between leaf Δ and TE for a range of sunflower genotypes grown in a glasshouse. Lambrides et al. (2004) surveyed 161 sunflower genotypes of diverse origin and found a large and unprecedented range of genetic variation for Δ (19.5-23.8‰). Their studies also showed a strong negative genetic correlation (rg) between TE and Δ (rg = -0.86, P<0.001) for fourteen inbreds that spanned the observed range for Δ, and five commercial hybrids when grown in a glasshouse. While these studies showed a strong relationship between TE and Δ, this needs to be verified using segregating populations. A study of the relationship between TE and Δ in segregating populations would provide valuable information on heritability and the manipulation of Δ in a breeding program. Lambrides et al. (2004) identified several sunflower inbred lines that differed significantly for TE. Two of these, HAR4 and SA52 have been crossed to develop a set of recombinant inbred lines (RILs). In the present study, 20 RILs selected from the HAR4 x SA52 population where included in a pot experiment and grown in the field under a rainout shelter to study the relationship between TE and Δ and their inheritance. F3, F5, and F7 segregants of HAR4xSA52 were also crossed to give hybrid combinations that were evaluated for grain yield under field conditions.

Materials and Methods

Rainout shelter experiment

A set of 20 HAR4 x SA52 recombinant inbred lines (RILs) that spanned a large range of Δ based on measurements of field grown plants were selected and evaluated for TE under a rainout shelter.

This experiment was conducted during March and April 2002 at the CSIRO Gatton Research Station (latitude 27 33' S, longitude 152 20' E ). Several seeds of each genotype were planted (March 20-21, 2002) in 25 litre “easi-lift”, black, polypropylene planter bags that had been covered in aluminium foil to reduce the heat load. Each bag was filled with 26 kg of a sandy alluvial loam and placed in a larger plastic bag to prevent drainage. Four replicate bags were planted per genotype, and bags were placed in rows one meter apart with bags 20 cm apart within rows. The planting arrangement was a randomized complete block design that also incorporated an alpha lattice.

The soil surface was covered with 800 grams (about 2 cm deep) of white alcathene (polyethylene) beads to minimise evaporation. The bags were watered to about nine-tenths field capacity by adding 4 litres of water. Nineteen days after emergence each bag was thinned to two plants per pot and weighed. For the next 2 weeks, by monitoring several bags with different plant size, water was added every two to three days in 594 ml or 1020 ml quantities using bottles with a narrow spout. Total water use (WUSE) was estimated by subtracting final bag weight (less above ground plant) from initial bag weight (less the weight of two seedlings removed at the thinning stage) and adding the amount of water that had been applied.

Plants were harvested 33-34 days after sowing when plants had produced 20 to 30 leaves. Plant height (HGT) was measured from the top of the apical meristem to the base of the stem. Plants were then cut at ground level, separated into leaves and stems and dried (80C for >48h) to determine leaf dry weight (LDW), stem dry weight (SDW) and above ground dry weight (AGDW= LDW + SDW). Leaf area (LA) was measured prior to drying. TE was calculated by dividing AGDW (minus initial plant weight at 19 days after sowing) by WUSE. Specific leaf weight (SLW) was calculated as LDW/LA.

After dry weights were recorded, leaves were ground through a 0.5 mm screen for carbon isotope analysis. All analyses were done by mass spectrometry at the Research School of Biological Sciences, Australian National University, Canberra. Δ is a measure of the 13C/12C ratio in plant material relative to the value of the same ratio in ambient air and was calculated according to Farquhar and Richards (1984). The units of Δ are per mil. (‰). Chlorophyll content of each genotype was estimated with a SPAD meter (Minolta, Osaka, Japan). Two SPAD measurements on one sunlit leaf per plant were made at the time of harvest.

Variance components were estimated with the COVTEST option of PROC MIXED (SAS 1990).

Estimates of narrow sense heritability on a genotype mean basis (h2) were calculated from variance components as the ratio of genetic variance to phenotypic variance.

Yield evaluations

Hybrid combinations were made by crossing sets of F3, F5 and F7 HAR4 x SA52 segregants, contrasting for Δ, to at least one of two genetic male tester lines named Pac-tester 1 and Pac-tester 2. In conjunction with Pacific Seeds, Toowoomba, the hybrids were yield tested using standard micro-plot techniques in 3-6 m single or two row plots. Dryland and irrigated trials were grown at the CSIRO Gatton Research Station, Australia in 2000 and dryland trials at Capella, Australia (latitude 23 05' S and longitude 148 01' E) in 2000 and 2001. The Capella site is in a major Australian sunflower growing region in central Queensland.

Results and Discussion

Lambrides et al. (2004) studied at set of unrelated sunflower inbred lines and observed a genetic correlation between TE and Δ of rg = -0.86. In the present study, a strong negative genetic correlation between TE and Δ was again observed, rg = -0.58 (Table 1). This is the first report describing the relationship between TE and Δ in a segregating population of sunflower. In addition, TE was strongly correlated to plant height at the final harvest (rg = 0.64) and TDW (rg = 0.58), and moderately correlated to SLW (rg = 0.46) and SPAD (rg = 0.21) but not leaf number (rg = 0.02). The lack of correlation between TE and leaf number contrasts with results of Lambrides et al. (2004) who observed a weak but positive genetic correlation.

Table 1. Genetic correlations among traits measured on twenty recombinant inbred lines selected from the population HAR4 x SA52 segregating for Δ and grown under a rainout shelter in Gatton, Queensland in 2002. Narrow sense heritability (h2) on a genotype mean basis is also shown.

Trait †

Trait

Genetic correlation

 

h2

h2

x

y

rg (xy)

se ‡ (rg)

x

y

TE

Δ

-0.58**

0.07

0.82

0.77

TE

SLW

0.46**

0.09

 

0.58

TE

SPAD

0.21*

0.09

 

0.94

TE

PHT

0.64**

0.06

 

0.88

TE

LNO

0.02 ns

0.10

 

0.82

TE

TDW

0.58**

0.07

 

0.83

* Significant at P <0.05, ** Significant at P <0.01, ns = not significant

† TE = transpiration efficiency (g DW kg/H20), Δ = 13C/12C discrimination (‰), SLW = specific leaf weight (g /m2), SPAD = a measure of chlorophyll content, PHT = plant height (cm), LNO = leaf number, TDW = total dry weight (g)

‡ se = standard error.

The associations of TE with SLW, SPAD and TDW suggests that photosynthetic capacity is a strong component of TE in sunflower. Stomatal conductance was not measured in this study but cannot be discounted as an important component of TE in sunflower based on the strong correlation between Δ and conductance observed by Lambrides et al. (2004).

Estimates of narrow sense heritability of TE and Δ were high (h2 = 0.82 and 0.77, respectively) suggesting that selection for these traits could be made in early generations of segregating populations. TE is relatively tedious to measure in large populations. Δ is much easier to measure but relatively expensive. SLW and SPAD could be used as less expensive surrogates of TE in an initial screen to discard germplasm in the early stages of a breeding program aimed at developing drought tolerant material.

Yield evaluations of hybrids contrasting for Δ showed that the low Δ pools of hybrids significantly out-yielded the high Δ pools in three dryland locations by 22-35% (Table 2). This indicates that selection for low Δ can significantly improve grain yield in dry environments that are characterised by yield levels of less than about 2 t/ha. While this yield advantage may not be fully achieved in commercial breeding programs, the data support our earlier computer simulations (data not shown) that yield improvements of 10-15% may be possible. Given that the ten year average yield of the commercial sunflower crop in Australia is 1 t/ha and largely limited by water supply, TE is therefore likely to be a valuable trait in breeding programs designed to develop drought tolerant hybrids.

Wheat varieties selected for low Δ have now been released in Australia and selection for TE looks to be equally promising for the development of drought tolerant sunflower parents and hybrids. Low Δ lines from the population HAR4 x SA52 have been distributed to private seed companies for further evaluation and for use as parents in their breeding programs.

Table 2. Grain yield and Δ for low vs high Δ pools of experimental top-cross hybrids made from (i) F3 segregates of the population HAR4 x SA52, evaluated in 2000 at Gatton and Capella, Australia and (ii) F5 segregates of the population HAR4 x SA52, evaluated in 2001 at Capella.

 

Gatton 2000
Irrigated

Gatton 2000
dryland

Capella 2000
dryland

Capella 2001
dryland

 

Yield
t/ha

Δ
(‰)

Yield
t/ha

Δ
(‰)

Yield
t/ha

Yield
t/ha

Δ
(‰)

High Δ pool

0.82
(100) †

20.7

0.48
(100)

22.0

1.01
(100)

1.58 (100)

20.4

Low Δ pool

0.89
(108)

20.3

0.65
(135)

21.6

1.37
(135)

1.93 (122)

20.1

Pr > F

0.52 ns

***

**

***

**

**

**

† percentages given in parentheses are relative to the high Δ group.

**P<0.01, *** P< 0.005

Acknowledgments

This research was supported by a grant from the GRDC (Grains Research and Development Corporation) to CSIRO. Field work support from Peter Harland is greatly appreciated. We also acknowledge the tester material and field trial support provided by Pacific Seeds Australia.

References

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Condon, A.G., R.A. Richards, and G.D. Farquhar. 1993. Relationships between carbon isotope discrimination, water use efficiency and transpiration efficiency for dryland wheat. Aust. J. Agric. Res. 44:1693-1711.

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Virgona, J.M., K.T. Hubick, H.M. Rawson, G.D. Farquhar, and R.W. Downes. 1990. Genotypic variation in transpiration efficiency, carbon-isotope discrimination and carbon allocation during early growth in sunflower. Aust. J. Plant Physiol. 17:207-214.

Wright, G.C., K.T. Hubick, G.D. Farquhar, and R.C. Nageswara Rao. 1993. Genetic and environmental variation in transpiration efficiency and its correlation with carbon isotope discrimination and specific leaf area in peanut. p. 247-267. In Ehleringer J.R. et al., (ed.) Stable isotopes and plant carbon-water relations. Academic Press, San Diego.

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