Fig. 8
Relative expression level of INV family members during low temperature in Citrus or its relative species. Two-month old seedlings from six Citrus species were treated on 4 °C for indicated time (0 h, 10 h, 24 h, 48 h and 72 h) and the leaves were collected for further analysis. Ptr, P. trifoliata; Ci, Citrus ichangensis; Cs, C. sinensis; Cg, C. grandis; Cl, C. limon and Fj, Fortunella japonica. Real-time quantitative (qRT-PCR) assay was performed using specific primers and PtrActin gene was utilized as a reference gene. The expression profiles of INV genes under cold stress were classified into the highly expressed genes (I group), the moderately expressed genes (III group), the slightly expressed (IV group) and no induction or down regulated genes (II group), which were colored by red, brown, gray and blue, respectively. The gene expression analysis for each treatment or each time point was repeated for at least four times. The relative gene expression was calculated using the 2-∆∆CT method relatively to the initial treatment time (0 h) that set as 1. The normalized values of the expression levels were converted into Log2 expression values and visualized by heatmap figure. Heatmap figures were constructed by TBtools based on default features, Euclidean distance method, and complete cluster method. Dark color refers to not determined values (n = 3)