BackgroundFragaria nilgerrensis(FN) provides a rich source of genetic variations for strawberry germplasm innovation. The color of strawberry fruits is a key factor affecting consumer preferences. However, the genetic basis of the fruit color formation inF. nilgerrensisand its interspecific hybrids has rarely been researched.ResultsIn this study, the fruit transcriptomes and flavonoid contents of FN (white skin; control) and its interspecific hybrids BF1 and BF2 (pale red skin) were compared. A total of 31 flavonoids were identified. Notably, two pelargonidin derivatives (pelargonidin-3-O-glucoside and pelargonidin-3-O-rutinoside) were revealed as potential key pigments for the coloration of BF1 and BF2 fruits. Additionally, dihydroflavonol 4-reductase (DFR) (LOC101293459 and LOC101293749) and anthocyanidin 3-O-glucosyltransferase (BZ1) (LOC101300000), which are crucial structural genes in the anthocyanidin biosynthetic pathway, had significantly up-regulated expression levels in the two FN interspecific hybrids. Moreover, most of the genes encoding transcription factors (e.g., MYB, WRKY, TCP, bHLH, AP2, and WD40) related to anthocyanin accumulation were differentially expressed. We also identified twoDFRgenes (LOC101293749 and LOC101293459) that were significantly correlated with members in bHLH, MYB, WD40, AP2, and bZIP families. Two chalcone synthase (CHS) (LOC101298162 and LOC101298456) and aBZ1gene (LOC101300000) were highly correlated with members in bHLH, WD40 and AP2 families.ConclusionsPelargonidin-3-O-glucoside and pelargonidin-3-O-rutinoside may be the key pigments contributing to the formation of pale red fruit skin.DFRandBZ1structural genes and some bHLH, MYB, WD40, AP2, and bZIP TF family members enhance the accumulation of two pelargonidin derivatives. This study provides important insights into the regulation of anthocyanidin biosynthesis in FN and its interspecific hybrids. The presented data may be relevant for improving strawberry fruit coloration via genetic engineering.Supplementary InformationThe online version contains supplementary material available at 10.1186/s12870-023-04361-1.