Mi Young Chung

Elimination of Chrysanthemum stunt viroid (CSVd) from meristem tip culture combined with prolonged cold treatment

Chrysanthemum production in Korea has recently been greatly affected by the spread of Chrysanthemum stunt viroid (CSVd) infection, necessitating the use of CSVd-free stocks to ensure successful chrysanthemum cultivation. We investigated the effects of low temperature (4°C), antiviral chemicals (ribavirin and amantadine) and a combination of these treatments on CSVd elimination by meristem tip cultures using plantlets that originated from CSVd-infected chrysanthemum ‘Ency’. Neither antiviral agents led to CSVd elimination, despite the suppression of meristem tip growth in a concentration dependent manner. However, the CSVd elimination rate increased up to 42.8% when meristem tips were excised after storage at 4°C for two months. The most effective results were obtained from a combination of low temperature for three months at 4°C followed by meristem tip culture on media containing 50 and 100 mg·L−1 ribavirin. These results suggest that antiviral agents can also be useful for CSVd elimination if their treatment is combined with prolonged periods of low temperature. This is the first report of eradication of viroids from spray type chrysanthemum bred in Korea.

Overexpression of snapdragon Delila ( Del ) gene in tobacco enhances anthocyanin accumulation and abiotic stress tolerance

BackgroundRosea1 (Ros1) and Delila (Del) co-expression controls anthocyanin accumulation in snapdragon flowers, while their overexpression in tomato strongly induces anthocyanin accumulation. However, little data exist on how Del expression alone influences anthocyanin accumulation.ResultsIn tobacco (Nicotiana tabacum ‘Xanthi’), Del expression enhanced leaf and flower anthocyanin production through regulating NtCHS, NtCHI, NtF3H, NtDFR, and NtANS transcript levels. Transgenic lines displayed different anthocyanin colors (e.g., pale red: T0-P, red: T0-R, and strong red: T0-S), resulting from varying levels of biosynthetic gene transcripts. Under salt stress, the T2 generation had higher total polyphenol content, radical (DPPH, ABTS) scavenging activities, antioxidant-related gene expression, as well as overall greater salt and drought tolerance than wild type (WT).ConclusionWe propose that Del overexpression elevates transcript levels of anthocyanin biosynthetic and antioxidant-related genes, leading to enhanced anthocyanin production and antioxidant activity. The resultant increase of anthocyanin and antioxidant activity improves abiotic stress tolerance.

Erratum to: Primary and secondary somatic embryogenesis in Chrysanthemum (Chrysanthemum morifolium) cv. ‘Baeksun’ and assessment of ploidy stability of somatic embryogenesis process by flow cytometry

We developed an efficient and simple system for inducing somatic embryogenesis and regenerating plantlets from petal explant of Chrysanthemum (Chrysanthemummorifolium) cv. ‘Baeksun’. Somatic embryogenesis was induced from petal explants on the Murashige and Skoog (MS) medium supplemented with 1.0 mg l−1 2,4-dichlorophenoxyacetic acid (2,4-D) and 3.0 mg l−1 6-benzyladenine (BA), yielding the highest mean number of embryos (56.3) per explant after 5 weeks of culture. We evaluated the effects of basal medium and various concentrations of sucrose on the proliferation of secondary somatic embryos. MS medium was observed to be more effective in promoting the proliferation of somatic embryos than half-strength Murashige and Skoog (1/2MS). In addition, 1 % sucrose was also found to be the best in induction of secondary embryogenesis. The highest germination rate (70 %) of the somatic embryos was observed on the MS medium containing 0.2 mg l−1 α-naphthalene acetic acid and 1 g l−1 activated charcoal (AC). Shoots elongated rapidly and roots developed well on hormone-free MS medium with 1 g l−1 AC and successfully acclimated in the greenhouse. Flow cytometric analysis of the primary somatic embryos, secondary somatic embryos, and the somatic embryo-obtained plants along with the parent grown in the greenhouse showed that they all had same identical peaks, indicating that there was no variation of ploidy level during the regeneration process. We expect that our report would be useful for micropropagation and Agrobacterium-mediated genetic transformation studies of this cultivar.

Elimination of chrysanthemum stunt viroid and chrysanthemum chlorotic mottle viroid from infected chrysanthemum by cryopreservation

Chrysanthemum morifolium ‘Borami’ and ‘Secret Pink’ showing symptoms of stunt disease caused by chrysanthemum stunt viroid (CSVd) and ‘Yellow Cap’ showing chlorotic mottle disease caused by chrysanthemum chlorotic mottle viroid (CChMVd) were confirmed to be infected by the respective viroids by using reverse transcription polymerase chain reaction (RT-PCR). Real-time PCR results showed that the viroid concentrations in the infected cultivars varied between the different regions of origin (Chilgok, Gumi, and Gyeongsan). We applied a cryopreservation protocol for elimination of CSVd from naturally infected ‘Borami’ collected from Gumi, showing the lowest concentration of CSVd, by varying several factors such as plant vitrification solutions (PVS2 and PVS3), duration of exposure to liquid nitrogen, shoot-tip size, and low-temperature treatment. The solution (PVS2) and low-temperature treatment were found to be critical factors determining the efficacy of viroid elimination. We optimized the protocol by combining of all resulted optimal factors and tested the applicability of the protocol in ‘Borami’ collected from Chilgok and Gyeongsan and in ‘Secret Pink’ from Chilgok, Gumi, and Gyeongsan, which displayed different viroid concentrations. We found that the elimination rates varied depending on the cultivar and region of origin. Similar results were observed when the protocol was applied to eliminate CChMVd from the ‘Yellow Cap’ collected from the same regions. Finally, we found that nested PCR is more reliable for viroid detection than RT-PCR. Overall, cryopreservation can be used to eliminate viroids from infected chrysanthemums; however, the efficacy depends on genotype and initial viroid concentration.

Sequence variation in SlMYB12 is associated with fruit peel color in pink tomato cultivars

The peel of pink-colored tomato is transparent due to the lack of accumulation of the flavonoid naringenin chalcone during ripening. A strong correlation was found between flavonoid expression and the function of SlMYB12, which is a transcriptional regulator of flavonoid biosynthesis. Thus, SlMYB12 is a strong candidate gene underlying the pink phenotype. Three allelic variants, a 603 bp deletion, a nucleotide substitution (C > T), and a 1 bp insertion (TG > TAG) in the SlMYB12 gene have been previously reported. We performed PCR genotyping based on these three allelic variations in 47 tomato cultivars displaying either a pink or red phenotype. However, the genotype did not match with the expected phenotype in one pink cultivar “Prime Alexander”. This cultivar was therefore self-pollinated to produce 20 progeny plants. To identify new mutations in SlMYB12, the sequence of genomic DNA and CDS were compared between the progeny 17 and the reference line, Heinz 1706. A novel G > T nucleotide substitution was found in the 2nd intron. This SNP leads to a deletion of 7 bp (GTAACAG) from the end of the 2nd exon, resulting in a premature stop codon. The presence of this SNP associates the pink phenotype with the genotype. This novel SNP will be useful as a genetic marker for marker-assisted breeding of pink tomato.