AbstractA significant proportion of hepatocellular carcinoma (HCC) is pathologically associated with hepatitis B virus (HBV) infection, followed by unsatisfied clinical outcomes. The increasing unmet need for HBV‐associated hepatocellular carcinoma (HBV‐HCC) treatment drives to deeper understand the role of the intricate immune microenvironment, tumor cell plasticity and dynamics of tumor evolution in HBV‐associated hepatic carcinogenesis. Thus, a comprehensive understanding of cross‐talk between HBV, host cells, and tumor microenvironment is of fundamental importance for identifying immune imbalance and heterogeneity in HBV‐HCC. Over the past 5 years, the application of single‐cell RNA sequencing (scRNA‐seq) in the understanding of heterogeneity and dynamics of immune cells, clonal evolution, and cancer stem cell (CSC) subsets of tumor cells has established a landscape for HBV‐HCC tumor ecosystem. Novel insights into anatomizing immune escape mechanisms and tumor drug resistance have remarkably facilitated the revolution of HBV‐HCC clinical treatment. Here, we provided a summary of HCC at single‐cell resolution and details on the basic workflow, limitations, and improvements of scRNA‐seq. The review highlights novel insights derived from scRNA‐seq on advances in the immune microenvironment and tumor heterogeneity of HBV‐HCC.