ABSTRACT Genetic improvement of manganese efficiency of crops demands a precise bioassay that avoids the confounding effect of seed manganese on the bioassay for the trait, for a proper comparison of the genotypes the seed manganese content must be similar. Variation in seed manganese can be minimized by the addition of manganese to individual seeds to be planted providing the effect of seed manganese content on early growth and development for the chosen genotype has been determined. It was hypothesized that growing the plants as mono-culms in very small pots under subclinical manganese deficiency would limit grain yield and yield attribute among genotypes, resulting in seed with less variation in manganese content across genotypes. The effect on yield and components of soil fertility ranging from University of California (UC) potting mix, fortified with Osmocote, to 100% manganese-deficient calcareous sand from Wangary, South Australia (Psamment) was determined for 20 bread wheat genotypes and breeding lines. It was observed that a mix of 70% UC and 30% Calcareous Wangary sand from South Australia in small pots minimized variation in grain yield and its components (number of tillers, number of seeds/head, weight of seed) and also lowered variation in seed manganese content among wheat genotypes. This was confirmed in pots of 70/30 UC-sand mix for five bread wheat cultivars, Worrakatta, Barunga, Schomburg, Excalibur, and Frame. The pot results from three replicates were compared with field results for the same genotypes grown at five randomly selected wheat-growing areas in South Australia: Paskeville, Geranium, Minnipa, Nunji, and Cummins. The variation in seed manganese content across genotypes was much less in pot grown seed. Aneuploid stocks (D-genome diosomic substitution lines) normally have highly variable seed size. Production of seed with similar manganese content was only feasible by growing the seed under multiple levels of soil manganese supply and after analysis selecting seeds with similar manganese content among the various manganese treatments. The effect of aneuploidy on seedling vigor, root growth, seed size and agronomic performance has been reported elsewhere. The effect on seed manganese content and the genetic analysis of micronutrient (manganese) efficiency were determined in the current study.