The nomilin and limonin content in citrus fruits of different varieties was determined at fruit growth and maturation stages by HPLC. The results showed that the two limonoids can be separated, identified, and quantified in citrus fruits within 10 min by the developed method. The method exhibited good precision, repeatability, stability, and recovery rate. The content of limonin and nomilin in most citrus fruits presented an increasing trend initially, and then decreased during fruit growth and maturation; a peak was observed at the young fruit or fruit expansion stage. The dropped fruits also contained some amount of limonoids, suggesting their industrial application. The variation and cluster analyses results revealed that the orange varieties contained the highest amount of limonoids at the mature stage. The results of this study will enable better use of citrus limonoids.

Methyl jasmonate (MeJA) is an airborne signaling phytohormone that can induce changes in endogenous jasmonates (JAs) and cause photosynthetic responses. However, the response of these two aspects of citrus plants at different MeJA concentrations is still unclear. Four MeJA concentrations were used in two citrus varieties, Huangguogan (C. reticulata × C. sinensis) and Shiranuhi [C. reticulata × (C. reticulata × C. sinensis)], to investigate the effects of MeJA dose on the endogenous JAs pathway and photosynthetic capacity. We observed that MeJA acted in a dosedependent manner, and its stimulation in citrus leaves showed a bidirectional character at different concentrations. This work demonstrates that MeJA at only a concentration of 2.2 mM or less contributed to the activation of magnesium protoporphyrin IX methyltransferase (ChlM, EC 2.1.1.11) and protochlorophyllide oxidoreductase (POR, EC 1.3.1.11) and the simultaneous accumulation of Chl a and Chl b, which in turn contributed to an improved photosynthetic capacity and PSII photochemistry efficiency of citrus. Meanwhile, the inhibition of endogenous JAs synthesis by exogenous MeJA was observed. This was achieved by reducing the ratio of monogalactosyl diacylglycerol (MGDG) to diagalactosyl diacylglycerol (DGDG) and inhibiting the activities of key enzymes in JAs synthesis, especially 12-oxo-phytodienoic acid reductase (OPR, EC 1.3.1.42). Another noteworthy finding is that there may exist a JA-independent pathway that could regulate 12-oxo-phytodienoic acid (OPDA) synthesis. This study jointly analyzed the internal hormone regulation mechanism and the external physiological response, as well as revealed the effects of exogenous MeJA on promoting the photosynthesis and inhibiting the endogenous JAs synthesis.

In this study we analysed the influence of drought stress on the leaf morphological characteristics, osmotic adjustment substances, antioxidant enzymes, and resistance-related photosynthetic physiological indices of Harumi tangor plants grafted onto Poncirus trifoliata (Pt), Citrus junos (Cj), and Citrus tangerine (Ct). The leaf relative water content and leaf area of the three rootstocks decreased with increasing drought stress, with the smallest decrease in Cj. The relative conductivity and malondialdehyde content increased with increasing drought stress. Proline, total soluble sugar, soluble protein, and activities of superoxide dismutase, ascorbate peroxidase, and catalase increased with drought stress but decreased under severe drought stress, with Cj exhibiting the greatest increase in enzyme activity. The net photosynthetic rate, stomatal conductance, transpiration rate, and chlorophyll a and b content were all lower than those of the control, whereas intercellular CO2 concentration increased with increasing drought stress. The initial fluorescence and maximal quantum yield of PSII were approximately equal for all rootstocks but increased with increasing drought stress severity. The combined analysis of physiological indicators, membership function, and principal components indicated that the drought resistance of grafted H. tangor decreased in the order Cj > Ct > Pt.

Citrus species are among the most economically important fruit crops. Physiological characteristics and molecular mechanisms associated with de-etiolation have been partially revealed. However, little is known about the mechanisms controlling the expression and function of genes associated with photosynthesis and chlorophyll biosynthesis in variegated citrus seedlings. The lower biomass, chlorophyll contents, and photosynthetic parameter values recorded for the variegated seedlings suggested that chlorophyll biosynthesis was partially inhibited. Additionally, roots of the variegated seedlings were longer than the roots of green seedlings. We obtained 567.07 million clean reads and 85.05 Gb of RNA-sequencing data, with more than 94.19% of the reads having a quality score of Q30 (sequencing error rate = 0.1%). Furthermore, we detected 4,786 and 7,007 differentially expressed genes (DEGs) between variegated and green Shiranuhi and Huangguogan seedlings. Thirty common pathways were differentially regulated, including pathways related to photosynthesis (GO: 0015979) and the chloroplast (GO: 0009507). Photosynthesis (44 and 63 DEGs), photosynthesis-antenna proteins (14 and 29 DEGs), and flavonoid biosynthesis (16 and 29 DEGs) pathways were the most common KEGG pathways detected in two analyzed libraries. Differences in the expression patterns of PsbQ, PetF, PetB, PsaA, PsaN, PsbP, PsaF, Cluster-2274.8338 (ZIP1), Cluster-2274.38688 (PTC52), and Cluster-2274.78784 might be responsible for the variegation in citrus seedlings. We completed a physiological- and transcriptome-level comparison of the Shiranuhi and Huangguogan cultivars that differ in terms of seedling variegation. We performed mRNA-seq analyses of variegated and green Shiranuhi and Huangguogan seedlings to explore the genes and regulatory pathways involved in the inhibition of chlorophyll biosynthesis and decreases in Chl a and Chl b contents. The candidate genes described herein should be investigated in greater detail to further characterize variegated citrus seedlings.

Ziyang xiangcheng (Citrus junos Sieb. ex Tanaka) (Cj) rootstock is effective in Citrus production; however, when Cj rootstock was used, sugar content in Kiyomi tangor fruit was significantly lower than that in the fruit produced using Poncirus trifoliata (L.) Raf. rootstock (Pt). Therefore, using K. tangor, we explored the cause of this difference, determining sugar accumulation, sucrose-metabolism enzyme activities, and gene expression. Before ripening, sugar content in fruits with Cj rootstock was significantly lower than that in fruits with Pt rootstock, due to low fructose and sucrose content. Sucrose phosphate synthase (SPS) activity of Pt was higher than that of Cj in the early growth stage (at 90–210 days after anthesis), however it was opposite at 240–300 days after anthesis. Additionally, neutral invertase (NI) activity of Pt was higher than that of Cj. Gene expression in Pt was higher than that in Cj, but is was essentially the same at maturity. SPS and NI activities and CitSPS1 expression were positively correlated with sucrose, fructose, and glucose content, but CSCW1 expression was negatively correlated with the sugars. Overall, the weak flavour of K. tangor fruit with Cj rootstock was regulated by the sucrose metabolism-related enzymes and gene expression.