<p>Ca²⁺ deficiency causes irreversible physiological disorders that markedly reduce the yield and quality of many horticultural crops, leading to substantial economic losses. Tip burn in Chinese cabbage (<i>Brassica rapa</i> ssp. <i>pekinensis</i>) is associated with Ca²⁺ deficiency; however, the underlying molecular mechanisms remain poorly understood. In this study, Ca²⁺ deficiency suppressed growth and impaired photosynthetic performance in Chinese cabbage seedlings. Under Ca²⁺ deficiency, cytosolic Ca²⁺ levels decreased, accompanied by excessive reactive oxygen species (ROS) accumulation in the leaves. Foliar application of dimethylthiourea (DMTU), an ROS scavenger, alleviated Ca²⁺ deficiency-induced tip burn by reducing ROS accumulation. RNA-seq analysis further revealed significant enrichment of the mitogen-activated protein kinase signaling and plant hormone signal transduction pathways during tip-burn development. Several differentially expressed genes associated with these pathways were identified as responsive to Ca²⁺ deficiency. Compared with normal Ca²⁺ and DMTU-rescued conditions, the <i>respiratory burst oxidase homolog D</i> (<i>BrRbohD-2</i>) gene was markedly upregulated under Ca²⁺ deficiency. Overexpression of <i>BrRbohD-2</i> increased ROS accumulation and aggravated tip burn in Chinese cabbage seedlings, whereas its suppression reduced ROS levels and alleviated tip burn. These findings provide important insights into the role of <i>BrRbohD-2</i> and the regulatory mechanisms underlying Ca²⁺ deficiency-related disorders in <i>Brassicaceae</i> crops.</p>