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Background  Esophageal squamous cell carcinoma (ESCC) cells are heterogeneous, easily develop radioresistance, and recur. Single-cell RNA-seq (scRNA-seq) is a next-generation sequencing method that can delineate diverse gene expression profiles of individual cells and mining their heterogeneous behaviors in response to irradiation. Our aim was using scRNA-seq to describe the difference between parental cells and cells that acquired radioresistance, and to investigate the dynamic changes of the transcriptome of cells in response to FIR.    Results  We sequenced ESCC cell lines KYSE180 with and without fractionated irradiation (FIR). A total of 218 scRNA-seq libraries were obtained from 88 cells exposed to 12 Gy (KYSE-180-12 Gy), 89 exposed to 30 Gy (KYSE-180-30 Gy), and 41 parental KYSE-180 cells not exposed to FIR. Dynamic gene expression patterns were determined by comprehensive consideration of genes and pathways. Biological experiments showed that KYSE-180 cells became radioresistant after FIR. PCA analysis of scRNA-seq data showed KYSE-180, KYSE-180-12 Gy and KYSE-180-30 Gy cells were discrete away from each other. Two sub-populations found in KYSE-180-12 Gy and only one remained in KYSE-180-30 Gy. This sub-population genes exposure to FIR through 12 Gy to 30 Gy were relevant to the PI3K-AKT pathway, pathways evading apoptosis, tumor cell migration, metastasis, or invasion pathways, and cell differentiation and proliferation pathways. We validated DEGs, such as  CFLAR ,  LAMA5 ,  ITGA6 ,  ITGB4 , and  SDC4  genes, in these five pathways as radioresistant genes in bulk cell RNA-seq data from ESCC tissue of a ESCC patient treated with radiotherapy and from KYSE-150 cell lines.    Conclusions  Our results delineated the divergent gene expression patterns of individual ESCC cells exposure to FIR, and displayed genes and pathways related to development of radioresistance.    Electronic supplementary material  The online version of this article (10.1186/s12864-019-5970-0) contains supplementary material, which is available to authorized users.

Single-cell RNA-seq of esophageal squamous cell carcinoma cell line with fractionated irradiation reveals radioresistant gene expression patterns