[Truncated abstract] Zinc deficiency in crops is the most widespread micronutrient deficiency, with about 50% of the cereal-growing areas worldwide containing low levels of plant-available Zn. Zinc plays multiple key roles in different metabolic and physiological processes; its deficiency in crops reduces not only grain yield, but also the nutritional quality of grains. Insufficient micronutrient intake, particularly Zn and Fe, afflicts over 3 billion people in the world, mainly in developing countries. Increasing the amount of Zn in food crops can contribute to improving the Zn status of people. Furthermore, Zn-dense seeds have agronomic benefits, resulting in greater seedling vigour, bigger root system and higher crop yield when sowed to soils with low plant-available Zn. Enhancing nutrient content and nutritional quality of crops for human nutrition is a global challenge currently, but it was mostly ignored during the breeding process in the past. There is a significant genotypic variation for seed Zn accumulation in several crops (including barley) which could be exploited in the breeding programs to produce genotypes with higher seed Zn concentration and content. However, the progress in Zn efficiency until now has mainly relied on conventional plant breeding approaches that have had limited success. Therefore, reliable alternative methods are required. Enhancing mineral nutrition through plant biotechnology may be a sustainable and beneficial approach in developing Zn-dense seeds in the staple crops. ... This DNA band was sequenced and converted into a simple sequence-specific PCR-based marker, which was designated as SZnR1 (seed Zn-regulator1). The developed marker is very easy to score, is inexpensive to run and amenable for a large number of plant samples. The successful development of SZnR1 molecular marker linked to chromosome region associated with seed Zn concentration and content using MFLP in this study illustrates the advantage of this technique over some other DNA fingerprinting methods used for identification of molecular markers for marker-assisted selection (MAS). In conclusion, the greater Zn efficiency of Sahara over Clipper under sufficient Zn supply may be attributed to its higher uptake of Zn. It appears that soil-based pot experiments under controlled condition may offer potential improvements over field experiments in screening for seed Zn accumulation. Shoot and seed Zn concentration and content can be used to diagnose the Zn statues of barley genotypes, and may be a useful selection criterion for Zn efficiency in large populations like doubled-haploid populations aimed at developing molecular markers for Zn efficiency. Identified QTLs influencing seed Zn concentration were repeatable in the field and glasshouse conditions, suggesting their robustness across environments as well as their value in marker-assisted selection. The developed PCR-based marker SZnR1 and other molecular markers associated with the QTLs on the short and long arms of chromosome 2H have the potential to be used for marker-assisted selection in breeding for Zn-dense seed in barley.