Comparative Transcriptomic and Physiological Analyses Reveal Key Factors for Interstocks to Improve Grafted Seedling Growth in Tangor.

Interstock is an important agronomic technique for regulating plant growth and fruit quality, and overcoming the incompatibility between rootstocks and scions; however, the underlying mechanisms remain largely unknown. In this study, the effects and regulatory mechanisms of tangor grafting, with and without interstocks, on the growth and development of scions were analyzed by combining morphology, physiology, anatomy and transcriptomics. Morphological and physiological analyses showed that interstocks ('Aiyuan 38' and 'Daya') significantly improved the growth of seedlings, effectively enhanced the foliar accumulation of chlorophyll and carotenoids, and increased the thickness of leaf tissues. Using 'Aiyuan 38' as the interstock, photosynthetic efficiency and starch content of citrus seedlings improved. Transcriptomics showed that genes related to photosynthesis and photosynthetic antenna proteins were upregulated in interstock-treated seedlings, with significant upregulation of photosystem PSI- and PSII-related genes. In addition, multiple key genes may be involved in plant hormone signaling, starch and sucrose metabolism, and transcriptional regulation. Taken together, these findings provide novel insights into the role of interstocks in regulating and contributing to the growth and development of grafted seedlings, and will further define and deploy candidate genes to explore the mechanisms of rootstock-interstock-scion interactions.

Transcriptome analysis of Harumi tangor fruits: Insights into interstock-mediated fruit quality.

Harumi tangor fruit with Ponkan as an interstock contains significantly higher levels of total soluble solids compared to Harumi tangor fruit cv.with no interstock. Transcriptome analysis of two graft combinations (Harumi/Hongjv (HP) and cv. cv.Harumi/Ponkan/Hongjv (HPP)) was conducted to identify the genes related to use of the Ponkan interstock. Soluble sugars and organic acids were also measured in the two graft combinations. The results showed that the contents of sucrose, glucose, and fructose were higher in the fruits of HPP than in those of HP; additionally, the titratable acid levels were lower in grafts with interstocks than in grafts without interstocks. Transcriptome analysis of HPP and HP citrus revealed that the interstock regulated auxin and ethylene signals, sugar and energy metabolism, and cell wall metabolism. Trend and Venn analyses suggested that genes related to carbohydrate-, energy-, and hormone-metabolic activities were more abundant in HPP plants than in HP plants during different periods. Moreover, weighted gene co-expression network analysis demonstrated that carbohydrates, hormones, cell wall, and transcription factors may be critical for interstock-mediated citrus fruit development and ripening. The contents of ethylene, auxin, cytokinin, transcription factors, starch, sucrose, glucose, fructose, and total sugar in HPP plants differed considerably than those in HP fruits. Interstocks may help to regulate the early ripening and quality of citrus fruit through the above-mentioned pathways. These findings provide information on the effects of interstock on plant growth and development.

Effects of Nitrogen Deficiency and Resupply on the Absorption of Mineral Nutrients by Tangor Cultivar 'Shiranuhi' (Citrus unshiu × C. sinensis) Grown in a Hydroponic System.

Tangor (Citrus unshiu × C. sinensis) cultivars obtained through interspecific hybridization have gained popularity in various countries, including South Korea. However, tangor has a relatively short history of cultivation and thus has been less well researched in terms of physiology and opping technology compared to mandarin. In addition, although tangor grows more vigorously than mandarin and thus has high nutrient demands and longer harvest time, it is more prone to various physiological disorders. Furthermore, the demand for nutrients could vary depending on the cultivars even within the same species. Thus, the current study examined the effects of nitrogen deficiency and resupply on the uptake of mineral nutrients using a specific cultivar 'Shiranuhi' as a case study. In this study, the tangor cultivar 'Shiranuhi' was cultivated in a hydroponics system, which allows the control of nutrient contents, and changes in mineral nutrient contents under nitrogen deficiency and nitrogen resupply were examined. Based on this, the results show the straightforward relationship between nitrogen and other mineral nutrients under a hydroponic system applying the cultivation of tangor. This implies that the hydroponics system can be applied to tangor cultivation and efficiently and widely expanded in Northeast Asia, and the income of growers will increase through the production of high-quality fruits.

Carotenoid biosynthesis and quality characteristics of new hybrids between tangor (Citrus reticulata x C. sinensis) cv. 'Murcott' and sweet orange (C. sinensis) cv. 'Pêra'.

Phenotypic characteristics, as well as the relation between carotenoid accumulation and gene expression during ripening were determined in fruits of five new hybrids between tangor cv. 'Murcott' and sweet orange cv. 'Pêra'. The genotypes were classified into the orange-like group, showing mainly epoxycarotenoids, oval fruit shape and yellowish color, or in the mandarin-like group, showing mainly ß-cryptoxanthin, flattened shape and deep-orange coloration; although some hybrids presented intermediate characteristics. The diversity in carotenoid composition of hybrids and genitors were mostly explained by patterns of gene expression. High carotenoid (250-426 µg/g dry weight [dw]) and ß-cryptoxanthin (81-125 µg/g dw) contents, observed in the mandarin-like group, were generally associated with high expression of upstream genes (GGPPS1, PSY, PDS). On the other hand, low expression/repression of these genes and high expression of downstream genes (BCHX and ZEP) were associated with low carotenoid (~158 µg/g dw) and ß-cryptoxanthin (5-22 µg/g dw) contents and epoxycarotenoid accumulation, as occurred in the orange-like group. Breeding experiments resulted in hybrids with outstanding higher carotenoid contents than both genitors (up to 426 µg/g dw versus 158-250 µg/g dw in genitors), which was attributed to transgressive segregation. Differences among genotypes have great impact on commercial fruit quality and potential health benefits, such as the provitamin A content.

Comparison of methodologies for conidia production by Alternaria alternata from citrus.

Conidia production is a problem in the study of Alternaria alternata from citrus. Thus, this study aimed to compare existing methodologies for conidial production of A. alternata isolated from Ponkan tangerine (2 isolates), Cravo lemon (1 isolate), Pêra orange (2 isolates) and Murcott tangor (1 isolate). The methodologies used were conidia production with 12 and 24 hours under white fluorescent light, evaluation with 24 and 48 hours after applying fungal mycelium stress technique, cold stress followed by injury of mycelium and evaluation with 24 hours, using healthy vegetable tissue and the use of black fluorescent near ultraviolet (NUV) lamp. Satisfactory result was obtained with A. alternata isolate from Murcott tangor, with the production of 2.8 × 10(5) conidia mL(-1), when fungal mycelium was stressed (Petri dish with 66.66% of fungi growth) and subsequently 24 h of growth. The use of white light (24 h) and black fluorescent NUV lamp also induced expressive conidia production by one isolate of Ponkan tangerine, which produced 17.2 × 10(5) and 10.1 × 10(5)conidia mL(-1) and another of Murcott tangor, which produced 13.9 × 10(5) and 10.1 × 10(5) conidia mL(-1), respectively. The remaining methodologies analyzed in this study were not able to induce conidia production in satisfactory quantity. The use of both mycelium stress technique and white light (24 h) and black fluorescent NUV lamp allowed the production of enough quantities of conidia to be used in vitro (detection of fungitoxic substances) and in vivo (pathogenicity test) assays, respectively.

Bioconverted Jeju Hallabong tangor (Citrus kiyomi × ponkan) peel extracts by cytolase enhance antioxidant and anti-inflammatory capacity in RAW 264.7 cells.

BACKGROUND/OBJECTIVES: Citrus and its peels have been used in Asian folk medicine due to abundant flavonoids and usage of citrus peels, which are byproducts from juice and/or jam processing, may be a good strategy. Therefore, the aim of this study was to examine antioxidant and anti-inflammatory effects of bioconversion of Jeju Hallabong tangor (Citrus kiyomi × ponkan; CKP) peels with cytolase (CKP-C) in RAW 264.7 cells. MATERIALS/METHODS: Glycosides of CKP were converted into aglycosides with cytolase treatment. RAW 264.7 cells were pre-treated with 0, 100, or 200 µg/ml of citrus peel extracts for 4 h, followed by stimulation with 1 µg/ml lipopolysaccharide (LPS) for 8 h. Cell viability, DPPH radical scavenging activity, nitric oxide (NO), and prostagladin E2 (PGE2) production were examined. Real time-PCR and western immunoblotting assay were performed for detection of mRNA and/or protein expression of pro-inflammatory mediators and cytokines, respectively. RESULTS: HPLC analysis showed that treatment of CKP with cytolase resulted in decreased flavanone rutinoside forms (narirutin and hesperidin) and increased flavanone aglycoside forms (naringenin and hesperetin). DPPH scavenging activities were observed in a dose-dependent manner for all of the citrus peel extracts and CKP-C was more potent than intact CKP. All of the citrus peel extracts decreased NO production by inducible nitric oxide synthase (iNOS) activity and PGE2 production by COX-2. Higher dose of CKP and all CKP-C groups significantly decreased mRNA and protein expression of LPS-stimulated iNOS. Only 200 µg/ml of CKP-C markedly decreased mRNA and protein expression of cyclooxygenase-2 in LPS-stimulated RAW 264.7 cells. Both 100 and 200 µg/ml of CKP-C notably inhibited mRNA levels of interleukin-1ß (IL-1ß) and IL-6, whereas 200 µg/ml CKP-C significantly inhibited mRNA levels of TNF-α. CONCLUSIONS: This result suggests that bioconversion of citrus peels with cytolase may enrich aglycoside flavanones of citrus peels and provide more potent functional food materials for prevention of chronic diseases attributable to oxidation and inflammation by increasing radical scavenging activity and suppressing pro-inflammatory mediators and cytokines.

Comparison of methodologies for conidia production by Alternaria alternata from citrus / Comparação de metodologias para produção de conídios por Alternaria alternata do citros

Conidia production is a problem in the study of Alternaria alternata from citrus. Thus, this study aimed to compare existing methodologies for conidial production of A. alternata isolated from Ponkan tangerine (2 isolates), Cravo lemon (1 isolate), Pêra orange (2 isolates) and Murcott tangor (1 isolate). The methodologies used were conidia production with 12 and 24 hours under white fluorescent light, evaluation with 24 and 48 hours after applying fungal mycelium stress technique, cold stress followed by injury of mycelium and evaluation with 24 hours, using healthy vegetable tissue and the use of black fluorescent near ultraviolet (NUV) lamp. Satisfactory result was obtained with A. alternata isolate from Murcott tangor, with the production of 2.8 x 10(5) conidia mL-1, when fungal mycelium was stressed (Petri dish with 66.66 percent of fungi growth) and subsequently 24 h of growth. The use of white light (24 h) and black fluorescent NUV lamp also induced expressive conidia production by one isolate of Ponkan tangerine, which produced 17.2 x 10(5) and 10.1 x 10(5) conidia mL-1 and another of Murcott tangor, which produced 13.9 x 10(5) and 10.1 x 10(5) conidia mL-1, respectively. The remaining methodologies analyzed in this study were not able to induce conidia production in satisfactory quantity. The use of both mycelium stress technique and white light (24 h) and black fluorescent NUV lamp allowed the production of enough quantities of conidia to be used in vitro (detection of fungitoxic substances)and in vivo (pathogenicity test)assays, respectively.

A produção de conídios consiste em problema no estudo de Alternaria alternata do citros. Assim, este estudo objetivou comparar metodologias existentes para a produção de conídios de A. alternata por dois isolados de tangerina Ponkan, um de limão Cravo, dois de laranja Pêra e um de tangor Murcott. As metodologias empregadas foram a produção de conídios com 12 e 24 horas sob luz branca, avaliação com 24 e 48 horas após estressamento do micélio do fungo, choque térmico com imediato estressamento do micélio e avaliação com 24 horas, produção de conídios pelo emprego de tecido vegetal sadio e o emprego de luz negra ultravioleta. Produção satisfatória de conídios foi obtida com o isolado de A. alternata de tangor Murcott, a qual foi de 2,8 x 10(5) conídios mL-1, mediante emprego da técnica de estressamento da colônia e cultivo do fungo por 24 horas. Os empregos de luz branca (24 h) e negra ultravioleta promoveram expressiva produção de conídios por um isolado de tangerina Ponkan, a qual foi de 17,2 x 10(5) e 10,1 x 10(5) conidios mL-1 e por outro de tangor Murcott, a qual foi de 13,9 x 10(5) e 10,1 x 10(5) conídios mL-1, respectivamente. As outras metodologias analisadas neste estudo não foram capazes de induzir a produção de conídios em quantidade satisfatória. Com o emprego das técnicas de estressamento do micélio e a utilização de luz branca (24 h) e negra ultravioleta, tornou-se possível obter quantidades de conídios suficientes para serem utilizadas em testes in vitro (detecção de substâncias fungitóxicas)e in vivo (testes de patogenicidade), respectivamente.