## 2 Result and Discussion
### Multivariate Analysis of Flower Volatiles during Development from Nine Cultivars  
To evaluate the genetic and developmental influences on flower volatiles, principle component analysis (PCA) and hierarchical cluster analysis (HCA) were performed. The PCA horizontal axis explained 19.78% of total variance and the vertical axis a further 15.68%, and three major groups, with the mandarin (2A, 2B, 2C), citron (1) and Pomelo (3) clearly separated, were observed (Figure 4), suggesting the existence of three major clusters with their subgroups as mentioned previously, and each subgroup was dominated by presence of specific major volatiles (Figure 4). More details were revealed by the HCA data, which also indicated that the nine cultivars could be clustered into three groups (Figure 5). Figure 4: Principle component analysis (PCA) of flower volatiles from nine citrus cultivars during blooming (abbreviated names of cultivars are presented in Table 1; each color represents one cultivar; while U, H and F represents unopened, half opened and fully opened flower stages). Note: each subgroup exhibited abundant volatiles. Figure 3: Ratio of major flower volatiles in either unopened (U) or fully opened (F) to half opened (H) flowers from nine citrus cultivars. Color code shown above the figure: red shows high, blue low. The blank cells indicate volatiles that were absent. **Figure 5.** Hierarchical cluster analysis based on flower volatiles during blooming for nine cultivars (abbreviated names of cultivars are as in Table 1, and **1** to **3** represent different cluster groups; while U represents unopened; H: half opened and F: fully opened flower blooming stages). Group **1** represented the citron, group **2** comprised four subgroups (**2A** Ponkan, **2B** sweet orange and Satsuma mandarin, and **2C** Satsuma mandarin and Huyou, and **2D** Yuhuanyou, Huyou and Qingjia), while group **3** represented pomelo and Huyou. Group 1 contains all citrons, all the samples yielded volatiles which were clearly differentiated from the other cultivars due to high amounts of limonene, which was the most abundant flower volatile in citrons (Tables 3-5). Group 2, comprising four subgroups, represented the volatiles profiling of Ponkan (2A), sweet oranges and Satsuma (F) (2B), Satsuma and Huyou (2C), as well as containing the volatiles of Yuhuanyou, Huyou and Qingjia (2D). Group 3 contains only pomelos and Huyou. These results are in agreement with the previous studies that the cultivars can be classified based on morphological and biochemical characteristics with citron, mandarin and pomelo identified as the only true citrus types [20,24,45]. The origin of other citrus, as hybrids between true citrus types, has been traced traditionally by molecular markers [20], and in this study, it was observed that the origin can be explored by volatile profiles as well. For example, Satsuma mandarin was clustered with sweet orange in subgroup 2B, rather than with Ponkan, which was in subgroup 2A. This supported the viewpoints of previous studies [46, 47, 48] where it has been reported that Satsuma mandarin is a hybrid and more closely related to sweet orange or pomelo rather than to mandarins, which is also consistent with the Tanaka classification [45], which classified Satsuma mandarin as separated from Ponkan. Flower volatile profiling data was also helpful for parentage analysis. For example, Huyou, which has been proposed to be a hybrid between a pomelo and a sour orange based on results from internal transcribed spacer (ITS) sequencing [40], was observed to be related to pomelos and a citrus belong to subgroup 2C, and the possibility that a sour orange could be clustered into this subgroup is worthy of further examination. Furthermore, it can also be concluded that volatile profiles of a hybrid could show different degree of contribution from its parents at different blooming stages.