As one of the top three tuber crops of the world, cassava is a staple food and feed crop for tropical and subtropical regions. Because of its high drought resistance and tolerance to nutrient deficiency, cassava is usually cultivated on hilly areas that are nutrient-poor. Nitrogen (N) is one of the significant factors affecting cassava yield. A double factorial (N level×genotypes) split-plot field experiment was conducted to analyze differences in yield and N accumulation of 25 cassava genotypes under low-N conditions to screen for cassava genotypes with high-N efficiency. The two-year field experiment showed that cassava yield and N accumulation are determined by specific genotypes, soil N levels, and year. Among these factors, soil N levels are the main factors that are responsible for differences in cassava yield. When yield and relative N accumulation under low-N conditions were used as screening markers, we identified an efficient and responsive genotype (SC10), and two inefficient and non-responsive genotypes (SC205 and GR5). The efficient and responsive genotype and the inefficient and non-responsive genotype can be used as study materials to further reveal the mechanisms for high-N efficiency in cassava.