The stability and yield of barley grain are affected by several factors, such as climatic conditions, fertilisation, and the different barley varieties. In a long-term experiment in Prague, Czech Republic, established in 1955, we analysed the weather trends and how weather, fertilisation (10 treatments in total), and different barley varieties affected grain yield and stability. A total of 44 seasons were evaluated. Trends in mean, minimum, and maximum temperatures from 1953 to 2023, as well as sunshine duration from 1961 to 2022, showed statistically significant increases. The trend for annual precipitation from 1953 to 2023 was not significant, but changes in precipitation were recorded via seasonal precipitation concentration indexes. The unfertilised Control and farmyard manure (FYM) provided the lowest mean yields. Mineral fertilisers (NPK) and FYM+NPK increased grain yield, ranging from 4.9 t ha−1 to 5.5 t ha−1. Three notable correlations between weather conditions and yields were observed: (1) June precipitation (r = 0.4), (2) minimal temperature in July (r = 0.3), and (3) sunshine duration in May (r = −0.5). According to the linear–plateau response model, the reasonable N dose is 55 kg ha−1, resulting in a mean yield of 6.7 t ha−1 for the contemporarily used barley variety Sebastián.