Suhyoung Park

Anthocyanin accumulation and expression of anthocyanin biosynthetic genes in radish (Raphanus sativus).

Radish [Raphanus sativus (Rs)] is an important dietary vegetable in Asian countries, especially China, Japan, and Korea. To elucidate the molecular mechanisms of anthocyanin accumulation in radish, the gene expression of enzymes directly involved in anthocyanin biosynthesis was analyzed. These genes include phenylalanine ammonia lyase (PAL), cinnamate 4-hydroxylase (C4H), 4-coumarate-CoA ligase (4CL), chalcone synthase (CHS), chalcone isomerase (CHI), flavanone 3-hydroxylase (F3H), dihydroflavonol reductase (DFR), and anthocyanidin synthase (ANS). RsDFR and RsANS were found to accumulate in the flesh or skin of two radish cultivars (Man Tang Hong and Hong Feng No.1). Radish skin contained higher CHS, CHI, and F3H transcript levels than radish flesh in all three cultivars. In the red radish, 16 anthocyanins were separated and identified by high-performance liquid chromatography (HPLC) and elctrospray ionization-tandem mass spectrometry (ESI-MS/MS). Some of them were acylated with coumaroyl, malonoyl, feruoyl, and caffeoyl moieties. Furthermore (-)-epicatechin and ferulic acid were also identified in the three cultivars.

Metabolic Profiling of Glucosinolates, Anthocyanins, Carotenoids, and Other Secondary Metabolites in Kohlrabi (Brassica oleracea var. gongylodes)

We profiled and quantified glucosinolates, anthocyanins, carotenoids, and other secondary metabolites in the skin and flesh of pale green and purple kohlrabis. Analysis of these distinct kohlrabis revealed the presence of 8 glucosinolates, 12 anthocyanins, 2 carotenoids, and 7 phenylpropanoids. Glucosinolate contents varied among the different parts and types of kohlrabi. Glucoerucin contents were 4-fold higher in the flesh of purple kohlrabi than those in the skin. Among the 12 anthocyanins, cyanidin 3-(feruloyl)(sinapoyl) diglucoside-5-glucoside levels were the highest. Carotenoid levels were much higher in the skins than the flesh of both types of kohlrabi. The levels of most phenylpropanoids were higher in purple kohlrabi than in pale green ones. trans-Cinnamic acid content was 12.7-fold higher in the flesh of purple kohlrabi than that in the pale green ones. Thus, the amounts of glucosinolates, anthocyanins, carotenoids, and phenylpropanoids varied widely, and the variations in these compounds between the two types of kohlrabi were significant.

Quantification of glucosinolates, anthocyanins, free amino acids, and vitamin C in inbred lines of cabbage (Brassica oleracea L.).

We profiled and quantified glucosinolates (GSLs), anthocyanins, free amino acids, and vitamin C metabolites in forty-five lines of green and red cabbages. Analysis of these distinct cabbages revealed the presence of 11 GSLs, 13 anthocyanins, 22 free amino acids, and vitamin C. GSL contents were varied amongst the different lines of cabbage. The total GSL content was mean 10.6 μmol/g DW, and sinigrin was the predominant GSL accounted mean 4.0 μmol/g DW (37.7% of the total) followed by glucoraphanin (1.9) and glucobrassicin (2.4). Amongst the 13 anthocyanins, cyanidin 3-(sinapoyl) diglucoside-5-glucoside levels were the highest. The amounts of total free amino acids in green cabbage lines ranged 365.9 mg/100g fresh weight (FW) to 1089.1mg/100g FW. Vitamin C levels were much higher in red cabbage line (129.9 mg/100g FW). Thus, the amounts of GSLs, anthocyanins, free amino acids, and vitamin C varied widely, and the variations in these compounds between the lines of cabbage were significant.

Simple and comprehensive SLA‐DQB1 genotyping using genomic PCR and direct sequencing

To enable the efficient analysis of a highly polymorphic swine major histocompatibility complex (MHC) class II gene, swine leukocyte antigen (SLA)-DQB1, we developed a simple and comprehensive high-resolution genotyping protocol. To obtain sufficient sequence information to design a set of common genotyping primers for SLA-DQB1, we cloned SLA-DQB1 introns 1 and 2 from 11 alleles with official four-digit allelic designations and sequenced the regions directly surrounding the SLA-DQB1 exon 2. Significant intronic nucleotide variations, including several deletions, were identified. Based on 733-bp assembled genomic sequences including introns 1 and 2 and exon 2 from 11 different alleles, a primer set was identified that allowed the ubiquitous amplification and analysis of the complete SLA-DQB1 exon 2 sequence. We then developed a method to directly sequence the amplified polymerase chain reaction (PCR) products without further experimental steps. We especially focused on avoiding superimposed peaks, which arose from the presence of allelic deletions, in the sequencing electropherogram of SLA-DQB1 heterozygous animals. The genotyping accuracy was evaluated by comparing the results of genomic sequence-based typing (GSBT) with those of other available methods, including cDNA sequence-based typing (SBT), low-resolution PCR typing with sequence-specific primers, allelic segregation analysis, and heterozygote simulation typing. In all cases, the results were consistent between SLA-DQB1 GSBT and previously reported methods or expected results. We applied it to genotype 350 animals from seven pig breeds. The observed level of heterozygosity from our genotyping was ∼51%, reflecting that a large portion of the animals were inbred miniature pigs. Among the seven pig breeds tested, the allelic diversity of SLA-DQB1 was highest in Berkshire pigs. In conclusion, we have developed a simple and effective SLA-DQB1 GSBT method by combining simple genomic DNA PCR and direct sequencing. Our new method may aid in the study of SLA diversity and disease resistance and susceptibility.

Metabolite profiles of glucosinolates in cabbage varieties (Brassica oleracea var. capitata) by season, color, and tissue position

Glucosinolates (GSLs) are sulfur-containing anionic secondary metabolites that are precursors of biologically active compounds, such as isothiocyanates, in Brassicaceae. The GSLs found in 10 cultivars of cabbage (Brassica oleracea var. capitata) popularly cultivated in Korea and China were identified and quantified. Three GSL classes (6 aliphatic, 1 aromatic, and 3 indolyl) were identified and quantified using high-performance liquid chromatography mass spectrometry (HPLC-MS) and HPLC. The concentration of these GSLs varied by season (spring- and fall-sown), leaf position (inside and outside), and cabbage color (green and red). The average total amounts of GSLs in the inner and outer sections of green and red cabbages ranged from 8.55–13.5 μmol∙g−1 dry weight. The spring-sown cabbages contained significantly higher GSL concentration (2.3–4.3 times higher) compared to their fall-sown counterparts. The inner sections of cabbages contained 1.1- to 1.8-fold greater GSL concentrations than the outer sections. These results indicate that an increase in temperature induces GSL accumulation in both green and red cabbages. The green cabbage cultivars contained significantly higher concentration of GSLs synthesized from homo-methionine, whereas the red cabbage cultivars had greater amounts of GSLs synthesized from dihomo-methionine, which suggests that the activities of enzymes involved in the elongation of homo-methionine are greater in red cabbages. The fall-sown red cabbages also contained 2- to 3-fold higher contents of GSLs synthesized from tryptophan compared to the spring-sown cabbages. In conclusion, the color of cabbage, tissue position of cabbage, temperature, and rainfall were all significantly correlated with the amount of GSL accumulation in cabbages.

Identification of three FLOWERING LOCUS C genes responsible for vernalization response in radish (Raphanus sativus L.)

Raphanus sativus L. is grown worldwide and used as fresh vegetables. In the Brassicaceae family, the FLOWERING LOCUS C (FLC) gene is a key regulator of flowering time and explains a large part of natural flowering time variation and the vernalization response. Here we report three FLC orthologous genes RsFLC1, RsFLC2, and RsFLC3 in R. sativus identified from the de novo assembled transcriptome. The sequences of three RsFLC genes have a high similarity to Arabidopsis FLC. Overexpression of each RsFLC gene in Arabidopsis induced late flowering, suggesting that every RsFLC gene functions as a floral repressor. All RsFLC genes were highly expressed in non-vernalized plants, whereas their expression levels significantly decreased by the vernalization treatment. Furthermore, the rate of decrease in their expression was proportional to the length of cold exposure. A significant level of sequence variation exists among RsFLC alleles derived from a variety of Raphanus cultivars, suggesting that RsFLC genes have diverged considerably but still retain essential functions.

Increase of phenolic compounds in new Chinese cabbage cultivar with red phenotype

A newly developed red Chinese cabbage (Brassica rapa ssp. campestris; CC) was analyzed to determine the changes in functional compounds, especially in phenolic compounds. In the red CC, cyanidin was the major anthocyanidin, with 4.11 mg·g−1 dry weight (dw), while pelargonidin and peonidin were minor components. The control CC, however, contained no anthocyanidins. In both CCs, four hydroxycinnamic acids were detected, and sinapic acid was the principal compound found. Hydroxycinnamic acids in the red CC were significantly more abundant than were those in the control, by a factor of 1.4–13.3. Quercetin was the principal flavonol in the red CC, with 2.7 mg·g−1 dw, whereas it was the flavonol with the lowest concentration in the control CC (0.5 mg·g−1 dw). Kaempferol and isorhamnetin concentrations were also increased in the red CC by 1.41- and 3.10-times, respectively. The results imply that, due to the common synthetic pathways of the phenolic compounds, the creation of a red phenotype from the presence of anthocyanidin increased the levels of flavonols and phenolic acids. Pectin, reducing sugar, and cellulose levels in the red CC indicate that the red CC could be preferred by consumers because of its better taste. Consequently, the new red CC, by accumulating significant amounts of the bioactive anthocyanidin, has significant value for human nutrition.

Identification and characterization of a novel mouse and human MOPT gene containing MORN-motif protein in testis.

A novel testis-derived membrane occupation and recognition nexus (MORN)-motif protein was identified in mouse testis (MOPT) by subtraction screening methods and found to be localized on chromosome 17E3, spanning approximately 7kb. Sequence analysis showed that MOPT contains 669 base pair nucleotides of open reading frame and the corresponding 79 amino acids. The protein is predicted to have theoretical molecular mass of 9000 Da and an expected isoelectric point of 5.8 and seems to have unique sequences except for MORN-motif domain. The transcript of MOPT is highly and specifically expressed in adult testis as well as skeletal muscle. Moreover, MOPT transcript and protein are confined mainly to round and elongated spermatids, except for a few individual dispersed spermatocytes, and increase in abundance at subsequent stages. MOPT first appeared in the proacrosomic vesicles of the early Golgi phase spermatids and was translocated from the head cap of elongated spermatid to the nucleus of mature spermatozoa at the final stage of spermiogenesis. Our study suggests that MOPT may play an important role in dynamic regulation of acrosome biogenesis during late spermiogenesis.

LED Lights Enhance Metabolites and Antioxidants in Chinese Cabbage and Kale

Light emitting diode (LED) lights play an important role in the plant physiology and alter the metabolites in a significant manner. Glucosinolates (GSLs), polyphenols, flavonoids and antioxidant properties of Chinese cabbage and kale cultivated in varying LED lights were investigated. Analysis revealed 7 aliphatic, 3 indolyl and 1 aromatic GSLs in Chinese cabbage and kale. The total GSL content ranged from 1.5-19.08 and 1.85-24.87 µmol/g DW, and glucobrassicanapin was the predominant GSL (3) in Chinese cabbage, whereas; sinigrin (3.49 µmol/g DW) was in kale. Blue and red LED lights produced significantly higher amount of GSLs in Chinese cabbage and kale respectively. Results revealed higher amount of total polyphenol (3.845 µg/mL) and total flavanoids (3.939 μg/mL) in Chinese cabbage. Chinese cabbage and kale showed significant antioxidant activities when compare with positive control, and the antioxidant assays were slightly correlated with total GSLs, polyphenols and flavanoids contents. The influence of LED lights on glucobrassicin in Chinese cabbage and kale should be studied extensively, because GSL is the precursor of indole-3-carbinol, a potent anticancer isothiocyanate.

Dormancy Associated Weedy Risk of the F1 Hybrid Resulted from Gene Flow from Oilseed Rape to Mustard

To assess the dormancy associated weedy risk of the F1 hybrid generated by hybridization between Brassica juncea (maternal) and Brassica napus (paternal), seed germination, dormancy and longevity were examined sequentially after seed harvest. The F1 hybrids exhibited the intermediate characteristics of their parents in seed germination and dormancy with relatively high dormancy rate of 41.1%. In summer, F1 hybrid seeds buried in the 3 cm soil exhibited greater viability (52.4%) than those in the soil surface with greater seed longevity (74.6%) than its maternal (63.3%) and paternal (33.7%) parents at 100 days of over-summering in soil. In winter, F1 seeds buried in the soil surface were more viable than those in the 3 cm soil with greater seed longevity (83.5%) than its maternal (39.0%) and paternal (71.7%) parents at 100 days of over-wintering in soil. Therefore, it is concluded that F1 hybrid resulted from gene flow from OSR to mustard has high seed dormancy and longevity during summer and winter, suggesting its weedy risk potential. Further studies are required to examine the reproductivity and fitness cost of F1 hybrid to make a clearer conclusion of its weedy risk.