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Background  Freezing injury, which is an important abiotic stress in horticultural crops, influences the growth and development and the production area of kiwifruit ( Actinidia  Lind1). Among  Actinidia  species,  Actinidia arguta  has excellent cold resistance, but knowledge relevant to molecular mechanisms is still limited. Understanding the mechanism underlying cold resistance in kiwifruit is important for breeding cold resistance.    Results  In our study, a population resulting from the cross of  A. arguta  ‘Ruby-3’ × ‘Kuilv’ male was generated for kiwifruit hardiness study, and 20 cold-tolerant and 20 cold-sensitive populations were selected from 492 populations according to their LT50. Then, we performed bulked segregant RNA-seq combined with single-molecule real-time sequencing to identify differentially expressed genes that provide cold hardiness. We found that the content of soluble sucrose and the activity of β-amylase were higher in the cold-tolerant population than in the cold-sensitive population. Upon − 30 °C low-temperature treatment, 126 differentially expressed genes were identify; the expression of 59 genes was up-regulated and that of 67 genes was down-regulated between the tolerant and sensitive pools, respectively. KEGG pathway analysis showed that the DEGs were primarily related to starch and sucrose metabolism, amino sugar and nucleotide sugar metabolism. Ten major key enzyme-encoding genes and two regulatory genes were up-regulated in the tolerant pool, and regulatory genes of the  CBF  pathway were found to be differentially expressed. In particular, a  14–3-3  gene was down-regulated and an  EBF  gene was up-regulated .  To validate the BSR-Seq results, 24 DEGs were assessed via qRT-PCR, and the results were consistent with those obtained by BSR-Seq.    Conclusion  Our research provides valuable insights into the mechanism related to cold resistance in  Actinidia  and identified potential genes that are important for cold resistance in kiwifruit.    Supplementary Information  The online version contains supplementary material available at 10.1186/s12864-021-07369-9.

BSR-Seq analysis provides insights into the cold stress response of Actinidia arguta F1 populations