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Global warming is contributing to extreme weather events such as abnormally colder weather, but there is lack of information on the freezing tolerance of citrus grown in Korea. Therefore, it is necessary to prevent damage from the cold in advance, by acquiring information on the critical temperature at which 50% injury (LT50) occurs for citrus varieties grown in Korea. Electrolyte leakage (EL) and tetrazolium chloride (TTC) analyses were performed to determine LT50 values and estimate the freezing tolerance for citrus varieties cultivated in Korea. We also investigated which factors such as genotype, environmental condition (e.g., in vitro, in vivo), cultivation condition, and wind speed affect the LT50 value in citrus. The LT50 value measured in the EL and TTC analyses of the ‘Haryejosaeng’ mandarin indicated it had better resistance to the cold than the citrus hybrids. The LT50 value in the in vitro experiments was lower than for the in vivo experiments, but there was little difference among the genotypes. Some factors, such as cultivation condition and wind speed, affected the LT50 values. The flavonoid levels, total polyphenol content, and antioxidant activity were affected by exposure to freezing temperatures, which was not related to the change in freezing tolerance level.

Anthocyanins, one of the major plant pigments, are responsible for the coloration of various plant tissues and organs, including fruits. They are known to have significant health benefits. Anthocyanin biosynthesis is influenced by environmental factors such as cold temperature and is transcriptionally regulated by MYB/bHLH/WD40 complexes composed of R2R3-MYB, basic helix-loop-helix (bHLH), and WD40-type transcription factors (TFs). The Ruby locus, which encodes a MYB TF, controls anthocyanin production in the fruit of blood orange varieties in a dominant manner. For the marker-assisted selection of citrus breeding lines with high anthocyanin production, we established PCR-based molecular markers for robust genotyping of Ruby locus alleles. Two dominant Ruby alleles, RD-1 and RD-2, were specifically detected in the blood orange varieties with high anthocyanin contents in their fruit flesh, but not in the varieties rich in lycopene. For functional analysis of Ruby alleles, we generated transgenic Arabidopsis expressing a functional wild-type R, a non-functional r-1, or a dominant RD-2 allele. Expression levels of flavonoid/anthocyanin biosynthetic genes and anthocyanin production were highly increased in the transgenic plants expressing the dominant Ruby allele by its retrotransposon-mediated native promoter under cold treatment, suggesting that the retrotransposon may contain cis-regulatory elements that respond to cold temperature. Our results also support that the MYB TF encoded by Ruby could be a key component of regulatory complexes that control flavonoid/anthocyanin biosynthesis in citrus.

Many citrus cultivars have the polyembryony trait that develops many nucellar embryos alongside a single zygotic embryo in an individual seed by sporophytic apomixis. This unique botanical trait hinders citrus breeding by genetic hybridization and affects breeding efficiency and cost. Techniques to efficiently identify nucellar and zygotic individuals in citrus are still very limited. For a systematic and targeted citrus breeding program, we collected 101 citrus genetic resources and determined their embryo types, which revealed 22 monoembryo, 54 polyembryo, and 25 mixed types. We also developed 17 simple sequence repeat (SSR) markers showing polymorphism among the genetic resources from the public resources and our own comparative genome analysis. Seventeen SSR markers detected a total of 181 alleles, ranging from 5 to 16 alleles per locus. The average polymorphism information content value was 0.67, ranging from 0.43 to 0.84. Genetic cluster analysis based on similarity matrices of alleles revealed that several genetic resources of the genus Citrus were fragmented and/or scattered throughout the entire dendrogram, not forming unique groups, due to frequent natural or intended genetic crossings. Application of these polymorphic SSR markers to F1 individuals derived from several genetic crosses using polyembryonic citrus cultivars as a female parent revealed that the polyembryony trait decreased the breeding efficiency due to the poor occurrence rate of zygotic individuals. Therefore, our results suggest that identification of nucellar and zygotic embryo-derived F1 individuals using SSR markers as a genotyping technology may be a powerful tool for establishing a systematic molecular breeding program in citrus.

세계적인 과수작물로서의 경제적 중요성에도 불구하고, 감귤 생산은 주로 자연교잡 실생이나 눈 돌연변이로부터의 선발 또는 단순 품종 도입 등을 통해 이루어지고 있는 실정이다. 긴 유년기, 다배성, 자가불화합성과 같은 감귤 고유의 식물학적 특성, 주요 형질들(병저항성, 수량성, 품질 등)의 QTL에 의한 조절 등은 전통...

Kiwifruit is a new fruit crop that was commercialized in the late 1970s. Recently, its cultivation and consumption have increased rapidly worldwide. Kiwifruit is a dioecious, deciduous, and climbing plant having fruit with hairs and various flesh colors and a variation in ploidy level; however, the industry consists of very simple cultivars or genotypes. The need for efficient cultivar improvement together with the evolutional and biological perspectives based on unique plant characteristics, have recently encouraged genome analysis and bioinformatics application. The draft genome sequence and chloroplast genome sequence of kiwifruit were released in 2013 and 2015, respectively; and gene annotation has been in progress. Recently, transcriptome analysis has shifted from previous ESTs analysis to the RNA-seq platform for intensive exploration of controlled genetic expression and gene discovery involved in fruit ascorbic acid biosynthesis, flesh coloration, maturation, and vine bacterial canker tolerance. For improving conventional breeding efficiency, molecular marker development and genetic linkage map construction have advanced from basic approaches using RFLP, RAPD, and AFLP to the development of NGS-based SSR and SNP markers linked to agronomically important traits and the construction of highly saturated linkage maps. However, genome and transcriptome studies have been limited in Korea. In the near future, kiwifruit genome and transcriptome studies are expected to translate to the practical application of molecular breeding.

Leaves of kiwifruit have long petiole and large size, so that they are easily damaged by strong wind. This study was performed to evaluate the effect of leaves loss by typhoon damage on fruit quality and yield and new cane growth with artificial defoliation. Five-year-old ‘Jecy Gold’ kiwifruit vines grown in a plastic house were used in this study. Artificial defoliation was done on August 23 (100 days after anthesis) at the different defoliation levels, such as 0 (control), 50, 75, and 100%. Cane regrowth by axillary bud burst, cane bark carbohydrate reserve, and fruit quality and yield were investigated. Severe defoliation at 75 and 100% level showed significantly higher axillary budburst than the control and the 50% defoliation. Especially, the 100% defoliation induced a few abnormal off-season flowering. Starch and sucrose contents were significantly declined during the first 2 weeks after defoliation at the 75 and 100% defoliation. However, other soluble carbohydrate reserves including fructose, glucose, and myo-inositol were not affected significantly. Fruit drop was promoted and the mean fruit weight, firmness, and soluble solid concentrations were lower by the defoliation. Our results indicated that the loss of leaves above 50% would induce a transient shortage of assimilate supply and reduce fruit quality and yield significantly.

Summary Desorption of catechins and caffeine from polyvinylpolypyrrolidone (PVPP) was comprehensively investigated. The result showed that caffeine could easily be desorbed from PVPP by the tested solvents except n-hexane, while catechins could only be thoroughly done by dimethyl sulphoxide (DMSO) and N, N-dimethylformamide (DMF). Excellent desorption efficiency of DMSO and DMF might be attributed to their high dipole moments and H-bond potentials. Addition of ethanol was recommended considering the elution efficiency and fluidity, but ethanol volume should be

The study was conducted to investigate the effect of fruit bearing type with leafy (LY) and leafless (LS) fruits on fruit growth, sugar, and acid characteristics in satsuma mandarin. Fruit growth including fruit length, diameter, shape index (diameter/length), weight, and size distribution was not different between LY and LS fruits at ripening time. Total soluble solids (TSS) concentration of the fruit juice increased and acidity decreased continuously and then TSS:acidity ratio increased with fruit development from 100 days after anthesis to ripening time. Soluble sugar was continuously increased, whereas organic acid decreased. Theses tendencies were related to the increase of sucrose and decline of citric acid, respectively. However, there was no effect of fruit bearing type on TSS, acidity, and TSS:acidity ratio, and soluble sugar and organic acid composition in fruit juice. Also, photosynthetic rate and chlorophyll index with SPAD value were not different between leaves adjacent to LY and LS fruits. The results indicated that fruit growth and sugar and acid characteristics were not affected by fruit bearing type with leafy and leafless fruits in satsuma mandarin.

Arabidopsis thaliana dynamin-like protein 1E (ADL1E) is known to regulate mitochondrial elongation. The adl1e mutant has no morphological phenotype, and the growth and photosynthetic activity of the mutant are similar to those of the wild type. Leaf O2 uptake, which is supported by mitochondrial activity in the dark, is increased 1.7-fold by mutation in adl1e gene. The ATP content in the dark of guard and mesophyll cell protoplasts (GCPs and MCPs, respectively) was 2.5- to 4-fold higher in GCPs of the mutant and the wild type, and increased upon the addition of glucose in both genotypes. Oligomycin, an inhibitor of mitochondrial ATPase, suppressed ATP synthesis in both GCPs and MCPS isolated from adl1e plants, indicating that mutant had higher mitochondrial activity. The stomatal apertures of mutant and wild-type plants were then analyzed in vitro. In the light, the stomata of both genotypes showed similar patterns of opening. However, in the dark response, the stomata of the adl1e mutant closed faster than did those of the wild type. Oligomycin severely inhibited dark-induced stomatal closure in both cell types. The results suggest that stomatal closure in the dark is governed by cytosolic ATP concentration, which is stimulated by mitochondrial activity.

Caffeine and catechins from the Yabukita, Yutakamidori, Saemidori, Okumidori, and Fushun varieties of tea leaves picked during different harvesting seasons from April to October were evaluated using HPLC. Total content of catechins increased greatly with the later harvesting time of tea leaves (i.e., picking the leaves in September versus in April) and decreased slightly after September. Yabukita tea leaves picked in August contained 43.1 mg% catechins including EGC, EC, ECG, and EGCG, with the ECGC levels constituting greater than 50% of those four compounds. Yutakamidori and Okumidori varieties picked in September contained the highest catechin values, at 43.6 mg% and 31.0 mg%, respectively. Fushun and Saemidori varieties contained lower catechin concentrations of 14.5 mg% (July) and 11.7 mg% (August) compared to other varieties. The EGCG levels gradually decreased in the late harvesting season, while levels of the other catechins, EC, EGC, and ECG, gradually increased. All varieties of green tea showed a gradual decrease in caffeine content toward the end of our harvesting efforts in October, with levels of 58～68 mg% in April and 28～57 mg% in October. Yabukita, Saemidori, and Okumidori varieties reached their highest caffeine levels in late spring/early summer, with Yabukita and Okumidori varieties reaching a high of 73.4% and 63.5% caffeine, respectively, in May, and Saemidori at 64.0% in June. In particular, Fushun still contained high caffeine of 66.8 mg% (September) during the late harvesting season.

Genetic transformation using a micro-cross section (MCS) technique was conducted to improve the carotenoid content in kiwifruit (Actinidia deliciosa cv. Hayward). The introduced carotenoid biosynthetic genes include geranylgeranyl diphosphate synthase (GGPS), phytoene desaturase (PDS), ζ-carotene desaturase (ZDS), β-carotene hydroxylase (CHX), and phytoene synthase (PSY). The transformed explants were selected on half-strength MS medium containing 0.001 mg l(-1) of 2,4-D and 0.1 mg l(-1) of zeatin, either 5 mg l(-1) hygromycin or 25 mg l(-1) kanamycin, and 500 mg l(-1) cefotaxime. The genomic PCR, genomic Southern blot analysis, and RT-PCR were performed to confirm the integration and expression of the transgenes. The transformation efficiencies of either kanamycin- or hygromycin-resistant shoots ranged from 2.9 to 22.1% depending on the target genes, and from 2.9 to 24.2% depending on the reporter genes. The selection efficiencies ranged from 66.7 to 100% for the target genes and from 95.8 to 100% for the reporter genes. Changes of carotenoid content in the several PCR-positive plants were determined by UPLC analysis. As a result, transgenic plants expressing either GGPS or PSY increased about 1.2- to 1.3-fold in lutein or β-carotene content compared to non-transgenic plants. Our results suggest that the Agrobacterium-mediated transformation efficiency of kiwifruit can be greatly increased by this MCS method and that the carotenoid biosynthetic pathway can be modified in kiwifruit by genetic transformation. Our results further suggest that GGPS and PSY genes could be major target genes to increase carotenoid contents in kiwifruit.

Kiwifruit shoots can be rapidly propagated through a micro-cross section (MCS) system we established here. Optimal culture conditions were determined for different expiant types, section sizes, and concentrations of inorganic salts and plant growth regulators. Rates of survival and callus formation were higher in half-strength MS salts than in full-strength MS media. Similar performance (i.e., survival and callus formation) was achieved with section sizes of either 800 μm or 1200 μm. Proliferation efficiency was greatest when expiants from stem tissue were cultured on 1/2 MS supplemented with 4.5x10-3 μM 2,4-dichlo-rophenoxyacetic acid and 4.6x10-1 μM zeatin. The number of shoots averaged 2.61 per expiant, representing an efficiency of 94%. RAPD analysis revealed that the regenerated plants from our MCS system were genetically stable. These results show that the culturing of micro-cross sections from stem tissue is a powerful method for kiwifruit propagation.

This study was conducted to determine the optimal transformation conditions during early gene transfer steps necessary to improve the efficiency of transformation mediated by Agrobacterium tumefaciens in Fuji apple plants. The use of 200 μM acetosyringone in the co-culture medium resulted in 4.7% transformation efficiency, compared with different concentrations of 0–400 μM. A 4-day co-culture period gave 1.9% transformation efficiency, compared with different co-culture periods of 1–5 days. There was no significant difference in transformation efficiency with different explant placement orientations on co-culture medium. A comparison of young leaves from plants cultured in rooting medium for 4 weeks with those cultured for 8 weeks showed that an 8-week culture period resulted in higher transformation efficiency. We therefore concluded that the efficiency of Fuji apple transformation depends on improving factors related to the gene transfer step.

In Fuji, the production of ethylene was increased with the addition of AgNO3 and inhibited with the addition of 10 μM aminoethoxyvinylglycine (AVG). The addition of 80 μM AgNO3 to transformed explants of ‘Fuji’ cultured on selection medium resulted in increased ethylene production (20 μl l−1) at 3 weeks. Under examining the effect of AgNO3 in ‘Fuji’, the 40 μM AgNO3 showed with higher 33.8% and 6.5% in the efficiency of regeneration and transformation. However, ethylene production in ‘Gala’ explants treated with 10μM AgNO3 (3 μl l−1) decreased after 2 weeks compared with the control (5 μl l−1). Although the regeneration efficiency of ‘Gala’ with 10 μM AgNO3 was higher (41.1%) than the control (20.1%), there was no significant difference in the transformation efficiency at the same concentration. Shoot regeneration of ‘Fuji’ and ‘Gala’ was completely inhibited with 10 μM AVG. These results suggest that the addition of AgNO3 affects the efficiency of Agrobacterium-mediated gene transfer in ‘Fuji’.

To develop a SCAR primer related to the hairy-fruit trait in the genusActinidia, we took a PCR-RAPD approach using arbitrary 10-mer primers. PCR with the UBC 376 primer generated specific fragments from three species with hairy fruit skin. Those fragments were then cloned to determine their nucleotide sequences. Two SCAR primers were designed from the UBC 376 primer and nucleotide sequences were obtained from the PCR fragments. A SCAR primer, OKC385, specifically amplified a 385-bp fragment from one clone ofActinidia eriantha, four ofActinidia chinensis, and four ofActinidia deliciosa. Deduced amino acid sequences of this fragment showed high sequence homology with plant cellulose synthases, which are involved in the biosynthesis of cellulose, a major cell wall component. The 385-bp fragment was specifically detected only in the seriesPerfectae C.F. Liang of sectionStellatae Li. This type has many hairs on the leaves, fruits, and stems, suggesting that the gene containing the PCR fragment is involved in hair formation in this phylogenetic group. Taken together, our results suggest that the SCAR primer, OKC385, can be used as a specific primer for early selection of the non-hair trait in breeding of the genusActinidia.

We isolated four cDNAs encoding different isoforms of NAD–SDH from Malus x domestica cv. Fuji. The derived amino acid sequences of MdSDH1 , MdSDH2 , MdSDH3 , and MdSDH4 have considerable homology with NAD–SDHs from human, rat, and sheep. MdSDH1 is almost identical with the previously reported apple NAD – SDH . MdSDH2 , MdSDH3 , and MdSDH4 are clearly distinguished from MdSDH1 based on the primary structure, showing 69–71% identity with MdSDH1 and 90–92% identity with each other. The expression of MdSDH2 , MdSDH3 , and MdSDH4 is restricted to sink tissues including young leaves, stems, roots, and maturing fruits, implying their relationship with the sink activity of apple tissues. Unlike the others, MdSDH1 is highly expressed in source leaves as well as young leaves, indicating that the expression of MdSDH1 is not restricted to sink tissues. During the fruit development, four SDH genes are highly expressed at the late stage, suggesting that they play an important role in the period of fruit maturation rather than in the early stage of fruit development. MdSDH2 and MdSDH3 fusion proteins expressed in Escherichia coli cells exhibit NAD–SDH activity in both direction, but not NADP-SDH activity. The present study indicates that MdSDH1 , MdSDH2 , MdSDH3 , and MdSDH4 encode the novel isoforms of NAD-dependent sorbitol dehydrogenase categorized to two independent phylogenetic groups.