Evaluating Heat Tolerance in Leaf Mustard (Brassica juncea) Using Cell Membrane Thermostability Across Multiple Environments with AMMI and GGE Biplot Analyses

Abstract

Heat stress is an increasing constraint on leaf mustard production (Brassica juncea); however, methods for screening heat tolerance and evaluating genotype stability across environments remain limited. This study established a rapid screening method for heat tolerance based on relative injury (RI, %), expressed as cell membrane thermostability (CMT) under heat treatments. A preliminary temperature-response experiment showed a sigmoidal increase in RI, with 45 °C providing the greatest discrimination among genotypes; therefore, it was selected as the evaluation temperature for heat tolerance. Six leaf mustard accessions from Myanmar and Japan were evaluated under five sowing conditions. The combined ANOVA revealed significant effects of genotype, environment and genotype-by-environment interaction, indicating that heat tolerance is genetically controlled but is also influenced by environmental conditions. Genotype stability across environments was evaluated using the additive main effects and multiplicative interaction (AMMI) and genotype plus genotype-by-environment (GGE) biplot. The analysis showed that JP256550 (M3) had the lowest and most favorable genotype selection index (GSI) for CMT, whereas Yamagata seisai and Miike takana, with the lowest AMMI stability value (ASV), showed the most stable CMT across environments. In contrast, JP256560 (M1) and JP256554 (M4) exhibited greater environmental sensitivity and specific adaptation. Different sowing times also accelerated plant development and flowering, with the Myanmar accessions bolting earlier than the Japanese accessions. The results indicated that CMT measured at 45 °C is an effective phenotyping tool for heat tolerance screening in combination with multi-environment stability analysis, providing a practical approach to identify stable genotypes and suggesting that M3 is a promising candidate for future heat-tolerance breeding.