Titanium dioxide nanoparticle (nTiO2) is one of the most produced nanoparticles worldwide. Its mechanism on crop development and performance is unclear as it is hard to predict their toxicity or benefit. Therefore, understanding the genetics of crop development under nTiO2 is a prerequisite for their applications in agriculture and crop improvement. Here, we aimed to examine the influnce of 300ppm nTiO2 on seed germination, seedling morphology, root-related traits in 121 worldwide spring barley (Hordeum vulgare L.) accessions. Results show that nTiO2 significantley affected all traits scored in this study. Response to nTiO2 treatment, clear wide natural variation among accesions was detected. Remarkably, 10 genotypes showed increased root length under nTiO2 at the seedling stage indicating that nTiO2 enhanced the root elongation. Genome-wide association scan (GWAS) was applied using 9K single