## 3 Results
### Effect of SA and MeJA on Carotenoid Content in Citrus Juice Sacs In Vitro  
The effects of SA and MeJA on the accumulation of the major carotenoids in the citrus juice sacs were also investigated. Although significant change in the total carotenoid content was not observed, the accumulation of individual carotenoid, such as \(\beta\)-cryptoxanthin and all-_trans_-violaxanthin, was affected by the SA treatment (Figure 2). Indeed, the content of all-_trans_-violaxanthin decreased, while that of \(\beta\)-cryptoxanthin, which accounts for about 30% of total carotenoid, increased after the SA treatment. On the contrary, the total carotenoid content was much lower than the control after the MeJA treatment. In particular, only the content of \(\beta\)-cryptoxanthin increased up to 0.24-fold with respect to the control; whilst the concentration of all-_trans_-violaxanthin, 9-_cis_-violaxanthin, and lutein significantly decreased (Figure 2). Effect of SA and MeJA on Expression of the Genes Related to Flavonoid and Carotenoid Accumulation in Citrus Juice Sacs  
In this study, the changes in the expression of the genes related to flavonoid and carotenoid metabolisms in response to SA and MeJA treatments were investigated by using real-time PCR. The results showed that the expression of _CitCHS1_ was downregulated, while the expression of _CitCHS2_, _CitCHI_, _CitUFIGT2_, and _CitS3H_ was significantly upregulated by the SA treatment (Figure 3). In the MeJA treatment, the expression of _CitCHS1_ and _CitF3H_ was downregulated, while the expression of _CitCHI_ and _CitUFIGT2_ was upregulated in the citrus juice sacs (Figure 3). Furthermore, the expression of five carotenoid biosynthetic genes (_CitPSY_, _CitPDS CitLCYe_, _CitLCYb1_ and _CitHYb_)  
Figure 1: Effect of salicylic acid (SA) and methyl jasmonate (MeJA) treatments on the flavonoid content in the juice sacs of Satsuma mandarin in vitro. The total flavonoid was calculated by summing riocictrin, narirutin, hesperidin, penicillin, and rhoifolia. Columns and bars represent the means and SE (\(n=3\)), respectively. Different letters indicate significant differences at the 5% level by [PERSON]’s HSD test. Figure 2: Effect of salicylic acid (SA) and methyl jasmonate (MeJA) treatments on the carotenoid content in the juice sacs of Satsuma mandarin in vitro. The total carotenoid was calculated by summing \(\beta\)-carotene, \(\beta\)-cryptoxanthin, all-_trans_-violaxanthin, 9-_cis_-violaxanthin, and lutein. Columns and bars represent the means and SE (\(n=3\)), respectively. Different letters indicate significant differences at the 5% level by [PERSON]’s HSD test. and one carotenoid catabolic gene (_CitNCED3_) was investigated in the citrus juice sacs. In the SA treatment, the expression of _CitPSY_, _CitPDS_, and _CitHYb_ was upregulated, while that of _CitNCED3_ was downregulated. Instead, in the MeJA treatment, the expression of _CitHYb_ was downregulated, and that of the other carotenoid metabolic genes was not significantly affected (Figure 4). Figure 4: Effect of salicylic acid (SA) and methyl jasmonate (MeJA) treatments on the expression of carotenoid metabolism related genes in the juice sacs of Satsuma mandarin in vitro. The mRNA levels were analyzed by TaqMan real-time RT-PCR, and the expression of 18S ribosomal RNA (rRNA) was used as a control to normalize the raw data. Columns and bars represent the means and SE (\(n=3\)), respectively. Different letters indicate significant differences at the 5% level by [PERSON]’s HSD test. PSY, phytoene synthase; PDS, 15-_cis_-phytoene desaturase; LCYc, lycopene epsilon-cyclase; LCYb1, lycopene beta-cyclase1; HYb, beta-carotene 3-hydroxylase; NCD3, 9-_cis_-epoxycarotenoid dioxygenase3. Figure 3: Effect of salicylic acid (SA) and methyl jasmonate (MeJA) treatments on the expression of flavonoid metabolism related genes in the juice sacs of Satsuma mandarin in vitro. The mRNA levels were analyzed by TaqMan real-time RT-PCR, and the expression of 18S ribosomal RNA (rRNA) was used as a control to normalize the raw data.