Cell-Based Phenotyping Reveals QTL for Membrane Potential Maintenance Associated with Hypoxia and Salinity Stress Tolerance in Barley

Abstract

Waterlogging and salinity are two major abiotic stresses that hamper crop production world-wide resulting in multibillion losses. Plant abiotic stress tolerance is conferred by many interrelated mechanisms. Amongst these, the cell's ability to maintain membrane potential (MP) is considered to be amongst the most crucial traits, a positive relationship between the ability of plants to maintain highly negative MP and its tolerance to both salinity and waterlogging stress. However, no attempts have been made to identify quantitative trait loci (QTL) conferring this trait. In this study, the microelectrode MIFE technique was used to measure the plasma membrane potential of epidermal root cells of 150 double haploid (DH) lines of barley (Hordeum vulgare L.) from a cross between a Chinese landrace TX9425 and Japanese malting cultivar Naso Nijo under hypoxic conditions. A major QTL for the MP in the epidermal root cells in hypoxia-exposed plants was identified. This QTL was located on 2H, at a similar position to the QTL for waterlogging and salinity tolerance reported in previous studies. Further analysis confirmed that MP showed a significant contribution to both waterlogging and salinity tolerance. The fact that the QTL for MP was controlled by a single major QTL illustrates the power of the single-cell phenotyping approach and opens prospects for fine mapping this QTL and thus being more effective in marker assisted selection.

Keywords: H+-ATPase; Hordeum vulgare; hypoxia; membrane potential; salinity tolerance; waterlogging tolerance.

FIGURE 1

Four steps of experimental procedure are illustrated. (A) Seedling is imbobilized in a vertical chamber and treated with hypoxia solution (N₂ bubbled 0.2% agar). (B) The vertical chamber is mounted in faraday cage for membrane potential measurements. (C) Electrode is positioned next to root epidermis. (D) Electrode is impaled into the root cell for membrane potential measurements.

FIGURE 2

The frequency distribution for membrane potential (MP) under hypoxia (0.2% agar) stress of DH lines derived from a cross of TX9425 and Naso Nijo.

FIGURE 3

Quantitative trait loci (QTL) for membrane potential, salt and waterlogging tolerances on 2HS. The figures related to salt and waterlogging tolerance incorporate data published by Xu et al. (2012). The full length of chromosome 2H is also displayed in this study.

FIGURE 4

Correlation between membrane potential and waterlogging tolerance scores (A) and between membrane potential and salt tolerance scores (B).

FIGURE 5

Quantitative trait loci associated with waterlogging tolerance (LOD values) on 2HS (A) and QTL associated with salt tolerance (LOD values) on 2HS (B). Black line: LOD value of original QTL; Red line: LOD value of QTL when membrane potential is used as a covariate.

FIGURE 6

Quantitative trait loci associated with membrane potential (LOD values) on 2HS. Black line: LOD value of original QTL; Red line: LOD value of QTL when waterlogging damage scores are used as a covariate; Green line: LOD value of QTL when salinity tolerance scores are used as a covariate.