Impacts of Deficit Irrigation on Photosynthetic Performance, Productivity and Nutritional Quality of Aeroponically Grown Tuscan Kale (Brassica oleracea L.) in a Tropical Greenhouse.

Tuscan kale was grown aeroponically with 5, 30 and 60 min nutrient spraying intervals (defined as 5 minNSIs, 30 minNSIs and 60 minNSIs). Four weeks after transplanting, some 5 minNSI plants were transferred to a 60 minNSI (5 minNSI → 60 minNSI) and 90 minNSI (5 minNSI → 90 minNSI) for one more week. Significantly lower light-saturated rates of photosynthesis and stomatal conductance were observed for plants grown with a 60 minNSI than with a 5 minNSI. However, all plants had similar internal CO2 concentrations and transpiration rates. Reduced light use efficiency but increased energy dissipation was observed in plants grown in a 60 minNSI. A higher nitrate concentration was observed in 60 minNSI plants compared to 5 minNSI and 30 minNSI plants, while all plants had similar concentrations of total reduced nitrogen, leaf soluble protein and Rubisco protein. Plants grown with prolonged NSIs (deficit irrigation) had lower biomass accumulation due to the inhibition of leaf initiation and expansion compared to 5 minNSIs. However, there was no substantial yield penalty in 5 minNSI → 60 minNSI plants. Enhancements in nutritional quality through deficit irrigation at pre-harvest were measured by proline and total soluble sugar. In conclusion, it is better to grow Tuscan kale with a 5 minNSI for four weeks followed by one week with a 60 minNSI before harvest to reduce water usage, yield penalty and enhance nutritional quality.

Brassica oleracea Var italica by-Products Prevent Lipid Accumulation and Cell Death in a Liver Cell Model of Lipid Toxicity.

Obesity, a rising concern in the Eastern world, encompasses several co-morbidities, namely non-alcoholic fatty liver disease (NAFLD). Potential natural-based interventions to decrease the burden of obesity complications are being investigated. Many of the edible parts of plants are not sold for consumption and end up as massive waste, losing nutritional potential. In fact, a sizeable amount of waste is generated within the different steps of the food supply chain, representing a massive loss of both plant material and natural resources. A good example is Brassica by-products (BBPs). The objective of this work was to investigate the effect of three different extracts from broccoli (Brassica oleracea var italica) by-products in an in vitro model of free fatty acid (FFA)-induced lipotoxicity using human hepatoma HepG2 cells. Broccoli leaf, stalk, and inflorescence extracts induced a dose-dependent decrease in the cell viability of HepG2 cells. However, the maximal non-lethal concentrations of leaves, stalks, and inflorescences (10 µg/mL) did not compromise mitochondrial function or neutral lipid accumulation in HepG2 cells. The extracts significantly decreased FFA-induced lipid accumulation in HepG2 cells either in a co-incubation or pre-incubation strategy. The broccoli extracts' capacity to prevent the FFA-induced decrease in catalase activity in HepG2 may explain the observed effects.

[Identification and expression analysis of NHX gene family in Chinese cabbage].

Na+/H+ antiporter (NHX) gene subfamily plays an important role in plant response to salt stress. In this study, we identified the NHX gene family members of Chinese cabbage and analyzed the expression patterns of BrNHXs gene in response to abiotic stresses such as high temperature, low temperature, drought and salt stress. The results showed that there were 9 members of the NHX gene family in Chinese cabbage, which were distributed on 6 chromosomes respectively. The number of amino acids was 513-1 154 aa, the relative molecular weight was 56 804.22-127 856.66 kDa, the isoelectric point was 5.35-7.68. Members of BrNHX gene family mainly existed in vacuoles, the gene structure is complete, and the number of exons is 11-22. The secondary structures of the proteins encoded by the NHX gene family in Chinese cabbage had alpha helix, beta turn and random coil, and the alpha helix occurred more frequently. Quantitative real-time PCR (qRT-PCR) analysis showed that the gene family members had different responses to high temperature, low temperature, drought and salt stress, and their expression levels differed significantly in different time periods. BrNHX02 and BrNHX09 had the most significant responses to these four stresses, and their expression levels were significantly up-regulated at 72 h after treatments, which could be used as candidate genes to further verify their functions.

Investigation of the Key Genes Associated with Anthocyanin Accumulation during Inner Leaf Reddening in Ornamental Kale (Brassica oleracea L. var. acephala).

Ornamental kale (Brassica oleracea L. var. acephala) is a popular decorative plant in late autumn and winter. However, only during low-temperature color-changed periods below rough 15 °C can the plant accumulate anthocyanins and exhibit a diverse array of foliar color patterns. In this study, we probed into the potential mechanism of inner leaf reddening in a red-leaf pure line of ornamental kale by physiological, metabolic, and transcriptomic analyses. Determination of anthocyanin contents in the uncolored new white leaves (S0), the light red leaves (S1) in the reddening period and the red leaves (S2) completing color change, and analysis of anthocyanin metabolites at stage S2, revealed that the coloring of red leaves was mainly attributed to the accumulation of cyanidins. We further used transcriptomic sequencing between the pairwise S0, S1, and S2 stages to identify 21 differentially expressed genes (DEGs) involved in anthocyanin biosynthesis, among which the expression level of 14 DEGs was positively correlated with anthocyanin accumulation, and 6 DEGs were negatively correlated with anthocyanin accumulation. A total of 89 co-expressed genes were screened out, from which three DEGs (BoCHI, Bo4CL3, and BoF3H) were identified as hub genes in co-expression DEGs network. BoDFR and BoCHI were the DEGs with the highest expressions at S2. Moreover, two co-expressed DEGs related to stress response (BoBBX17 and BoCOR47) also exhibited upregulated expressions and positive correlations with anthocyanin accumulation. A deep dive into the underlying regulatory network of anthocyanin accumulation comprising these six upregulated DEGs from S0 to S2 was performed via trend, correlation, and differentially co-expression analysis. This study uncovered the DEGs expression profiles associated with anthocyanin accumulation during ornamental kale inner leaf reddening, which provided a basis for further dissecting the molecular mechanisms of leaf color characteristic change in ornamental kale at low temperatures.

Melatonin Delays Postharvest Senescence through Suppressing the Inhibition of BrERF2/BrERF109 on Flavonoid Biosynthesis in Flowering Chinese Cabbage.

Flowering Chinese cabbage is prone to withering, yellowing and deterioration after harvest. Melatonin plays a remarkable role in delaying leaf senescence and increasing flavonoid biosynthesis. However, the underlying molecular mechanisms of melatonin procrastinating postharvest senescence by regulating flavonoid biosynthesis remain largely unknown. In this study, melatonin could promote flavonoid accumulation and delay the postharvest senescence of flowering Chinese cabbage. Surprisingly, we observed that BrFLS1 and BrFLS3.2 were core contributors in flavonoid biosynthesis, and BrERF2 and BrERF109 were crucial ethylene response factors (ERFs) through the virus-induced gene silencing (VIGS) technique, which is involved in regulating the postharvest senescence under melatonin treatment. Furthermore, yeast one-hybrid (Y1H), dual luciferase (LUC), and ß-glucuronidase (GUS) tissue staining experiments demonstrated that BrERF2/BrERF109 negatively regulated the transcripts of BrFLS1 and BrFLS3.2 by directly binding to their promoters, respectively. Silencing BrERF2/BrERF109 significantly upregulated the transcripts of BrFLS1 and BrFLS3.2, promoting flavonoid accumulation, and postponing the leaf senescence. Our results provided a new insight into the molecular regulatory network of melatonin delaying leaf senescence and initially ascertained that melatonin promoted flavonoid accumulation by suppressing the inhibition of BrERF2/BrERF109 on the transcripts of BrFLS1 and BrFLS3.2, which led to delaying the leaf senescence of postharvest flowering Chinese cabbage.

Do differentially charged nanoplastics affect imidacloprid uptake, translocation, and metabolism in Chinese flowering cabbage?

Micro(nano)plastics are ubiquitous in the environment. Among the microplastics, imidacloprid (IMI) concentration has been increasing in some intensive agricultural regions, thus receiving increased attention. However, only a few studies have investigated the interaction of nanoplastics (polystyrene (PS)) and IMI in vegetable crops. We studied the effects of positively (PS-NH2) and negatively (PS-COOH) charged nanoplastics on the uptake, translocation, and degradation of IMI in Chinese flowering cabbage grown in Hoagland solution for 28 days. PS-NH2 co-exposure with IMI inhibited plant growth, resulting in decreased plant weight, height, and root length. Translocation of IMI from the roots to the shoots was significantly lower in the presence of PS-NH2, whereas PS-COOH accelerated the accumulation and translocation of IMI in plants, thus potentially affecting IMI metabolism in plants. Notably, IMI-NTG and 5-OH-IMI were the two dominant metabolites. PS-NH2 co-exposure with IMI induced significant oxidation stress and considerably affected the activities of superoxide dismutase (SOD) and peroxidase (POD), indicating that the antioxidant defense system was the main mechanism for reducing oxidative damage. Notably, both positively and negatively charged nanoplastics can accumulate in Chinese flowering cabbage. Plants in the PS-COOH alone treatment group had the highest concentration of nanoplastics in both roots and shoots. The accumulation of nanoplastics, IMI, and its metabolites in plants raises concerns about their combined potential toxicity because it compromises food safety.

Identification of the BcLEA Gene Family and Functional Analysis of the BcLEA73 Gene in Wucai (Brassica campestris L.).

Late embryogenesis abundant (LEA) proteins are important developmental proteins in the response of plants to abiotic stress. In our previous study, BcLEA73 was differentially expressed under low-temperature stress. Herein, we combined bioinformatics analysis, subcellular localization, expression assays, and stress experiments (including salt, drought, and osmotic stress) to identify and analyze the BcLEA gene family. Gene cloning and functional analysis of BcLEA73 were performed in tobacco and Arabidopsis. Based on the sequence homology and the available conservative motif, 82 BrLEA gene family members were identified and were divided into eight subfamilies in the genome-wide database of Chinese cabbage. The analysis showed that the BrLEA73 gene was located on chromosome A09 and belonged to the LEA_6 subfamily. Quantitative real-time PCR analysis indicated that the BcLEA genes were differentially expressed to varying degrees in the roots, stems, leaves, and petioles of Wucai. The overexpressed BcLEA73 transgenic plants exhibited no significant differences in root length and seed germination rates compared to the wild-type (WT) plants under control conditions. Under salt and osmotic stress treatment, the root length and seed germination rates of the BcLEA73-OE strain were significantly greater than those of WT plants. Under salt stress, the total antioxidant capacity (T-AOC) of the BcLEA73-OE lines increased significantly, and the relative conductivity, (REL), hydrogen peroxide (H2O2) content, and superoxide anion (O2-) production rate decreased significantly. Under drought treatment, the survival rate of the BcLEA73-OE lines was significantly higher than that of WT plants. These results showed that the BcLEA73 gene of Wucai functions in enhancing the tolerance of plants to salt, drought, and osmotic stress. This study provides a theoretical basis to explore the relevant functions of the BcLEA gene family members of Wucai.

Glucosinolates and Cytotoxic Activity of Collard Volatiles Obtained Using Microwave-Assisted Extraction.

Glucosinolates (GSLs) in Brassica oleracea L. convar. acephala var. viridis (collard) flower, leaf, stem, and root were analyzed qualitatively and quantitatively via their desulfo-counterparts using UHPLC-DAD-MS/MS. Twelve GSLs were identified, including Met-derived GSLs (sinigrin, glucoibervirin, glucoerucin, glucoiberin, glucoraphanin, progoitrin), Trp-derived GSLs (4-hydroxyglucobrassicin, glucobrassicin, 4-methoxyglucobrassicin, and neoglucobrassicin), and Phe-derived GSLs (glucotropaeolin and gluconasturtiin). Total GSL content was highest in the root, having 63.40 µmol/g dried weight (DW), with gluconasturtiin (34.02 µmol/g DW) as the major GSL, followed by sinigrin and glucoibervirin (12.43 and 7.65 µmol/g DW, respectively). Total GSL contents in the flower, leaf, and stem were lower than in root, having 6.27, 2.64, and 1.84 µmol/g DW, respectively, with Trp and/or Met-derived GSLs as the predominant ones. GSL breakdown products were obtained via microwave hydrodiffusion and gravity (MHG) and volatile breakdown products were analyzed using GC-MS techniques. Volatile isolates were tested for their cytotoxic activity using MTT assay. MHG volatile extract from the root demonstrated the best cytotoxic activity against human bladder cancer cell line T24 and breast cancer cell line MDA-MB-231 during an incubation time of 72 h (IC50 21.58, and 11.62 µg/mL, respectively). The activity of the root extract can be attributed to its major volatile, 2-phenylethyl isothiocyanate (gluconasturtiin breakdown product).

Insights into the loss of glucoraphanin in post-harvested broccoli--Possible involvement of the declined supply capacity of sulfur donor.

The loss of characteristic nutrient glucoraphanin during the shelf life seriously affects the nutritional quality of broccoli. Here, we monitored the changes in the levels of sulfur donors (cysteine and glutathione) required for glucoraphanin biosynthesis. Similar to glucoraphanin, cysteine content decreased sharply. Continuous down-regulation of BoCysK1 and BoCysK2 genes encoding cysteine synthase might account for cysteine loss. Contrarily, glutathione content accumulated steadily, which might owe to the up-regulation of biosynthetic gene (BoEC1). Additionally, the change of malondialdehyde content was positively correlated with glutathione, implying that oxidative stress might stimulate glutathione accumulation. Nevertheless, the expression of BoGSTF11 gene encoding glutathione S-transferases was down-regulated, which blocked the supply of glutathione. The increase in the content of raphanusamic acid (degradation product) indicated that insufficient supply of sulfur donors not only could constrain the biosynthesis of glucoraphanin but also triggered its degradation.

Exogenous brassinosteroid alleviates calcium deficiency induced tip-burn by regulating calcium transport in Brassica rapa L. ssp. pekinensis.

Mini Chinese cabbage (Brassica rapa L. ssp. Pekinensis) plays an important role in the supply of summer vegetables on the plateau in western China. In recent years, tip-burn has seriously affected the yield, quality and commodity value of mini Chinese cabbage. Calcium (Ca2+) deficiency is a key inducer of tip-burn. As a new type plant hormone, brassinolide (BR) is involved in regulating a variety of biotic and abiotic stresses. To explore the alleviation role of BR in tip-burn caused by Ca2+ deficiency, a hydroponic experiment was conducted to study the relationship between BR and Ca2+ absorption and transport. The results showed that foliar spraying with 0.5 µM BR significantly reduced tip-burn incidence rate and disease index of mini Chinese cabbage caused by Ca2+ deficiency. Moreover, the dynamic monitoring results of tip-burn incidence rate showed that the value reached the highest on the ninth day after treatment. BR promoted the Ca2+ transport from roots to shoots and from outer leaves to inner leaves by increasing the activities of Ca2+-ATPase and H+-ATPase as well as the total ATP content, which provided power for Ca2+ transport. In addition, exogenous BR upregulated the relative expression levels of BrACA4, BrACA11, BrECA1, BrECA3, BrECA4, BrCAX1, BrCAS and BrCRT2, whereas Ca2+ deficiency induced down-regulation. In conclusion, exogenous BR can alleviate the Ca2+-deficiency induced tip-burn of mini Chinese cabbage by promoting the transport and distribution of Ca2+.

The responses of organic acid production and microbial community to different carbon source additions during the anaerobic fermentation of Chinese cabbage waste.

The effects of glucose, fructose, sucrose and molasses on organic acid levels, protein degradation, nutrient preservation and bacteriome were studied during the anaerobic fermentation of Chinese cabbage waste. The results showed that fructose and molasses additions caused a significant (p < 0.05) increase in lactic acid production (82.16-89.79 %), acetic acid production (175.41-196.93 %), ammonia nitrogen formation (15.93-37.43 %) and reduction of neutral detergent fiber level (8.17-15.87 %). However, few positive effects of glucose and sucrose additions were found on organic acid production. Furthermore, carbon source additions enriched (p < 0.05) the acid-producing bacteria, such as Lactobacillus paralimentarius and Lactobacillus heilongjiangensis, upregulated (p < 0.05) the pathways of carbohydrate and lipid metabolisms and reduced (p < 0.05) the abundances of Lactobacillus buchneri and Escherichia coli and bacteria that were mobile elements-contained and stress-tolerant. Collectively, fructose and molasses additions enhanced the recycling of Chinese cabbage waste by anaerobic fermentation, in which the desired products are organic acids.

Evidence of glucosinolates translocation from inflorescences to stems during postharvest storage of broccoli.

Broccoli is a vegetable appreciated by consumers for its nutritional properties, particularly for its high glucosinolate (GLS) content. However, broccoli shows a high rate of senescence during postharvest and the GLS content in inflorescences decreases sharply. Usually, postharvest studies on broccoli focus on inflorescences, ignoring the other tissues harvested such as the stems and main stalk. In this work, GLS metabolism in whole heads of broccoli (including inflorescences, small stems and stalk) was analysed during postharvest senescence. The content of GLS content, expression of GLS metabolic genes, and expression of GLS transport-associated genes were measured in the three parts of harvested broccoli. A marked decrease in the content of all GLSs was detected in inflorescences, but an increase in the stems and stalk. Also, decreased expressions of GLS biosynthesis and degradation genes were detected in all tissues analysed. On the other hand, an increase in the expression of one of the genes involved in GLS transport was observed. These results suggest that GLSs would be transported from inflorescences to stems during postharvest senescence. From a commercial point of view, broccoli stems are usually discarded and not used as food. However, the accumulation of GLSs in the stems is an important factor to consider when contemplating potential commercial use of this part of the plant.

The effects of short term blue light treatment on promoting nutrition value in Chinese cabbage.

Chinese cabbage is a nutrients-rich vegetable with diverse leaf colors. Here, we used widely-targeted metabolomics technology to study the metabolic responses of three Chinese cabbage varieties with representative leaf colors after blue light treatment. The inner leaf color of orange varieties 20S530 and 15S1094 changed from yellow to golden yellow, while no visible color change occurred in the common variety 14S23 after the treatment. A total of 844 metabolites were measured from the leaf samples of these three varieties in a time course study after short term blue light treatment, with kaempferol-4'-O-glucoside, isoquercitrin, hyperin, arbutin, sulforaphane as enriched nutritional metabolites. Orange Chinese cabbage varieties showed additional nutrition enhancement after the treatment. This study is the first to explore the global metabolic responses of Chinese cabbage after blue light treatment, and our findings provided valuable insights on how to effectively use lighting conditions to enhance specific groups of nutrients in vegetables.

Pollen Coat Proteomes of Arabidopsis thaliana, Arabidopsis lyrata, and Brassica oleracea Reveal Remarkable Diversity of Small Cysteine-Rich Proteins at the Pollen-Stigma Interface.

The pollen coat is the outermost domain of the pollen grain and is largely derived from the anther tapetum, which is a secretory tissue that degenerates late in pollen development. By being localised at the interface of the pollen-stigma interaction, the pollen coat plays a central role in mediating early pollination events, including molecular recognition. Amongst species of the Brassicaceae, a growing body of data has revealed that the pollen coat carries a range of proteins, with a number of small cysteine-rich proteins (CRPs) being identified as important regulators of the pollen-stigma interaction. By utilising a state-of-the-art liquid chromatography/tandem mass spectrometry (LC-MS/MS) approach, rich pollen coat proteomic profiles were obtained for Arabidopsis thaliana, Arabidopsis lyrata, and Brassica oleracea, which greatly extended previous datasets. All three proteomes revealed a strikingly large number of small CRPs that were not previously reported as pollen coat components. The profiling also uncovered a wide range of other protein families, many of which were enriched in the pollen coat proteomes and had functions associated with signal transduction, cell walls, lipid metabolism and defence. These proteomes provide an excellent source of molecular targets for future investigations into the pollen-stigma interaction and its potential evolutionary links to plant-pathogen interactions.

Inactivation of RPX1 in Arabidopsis confers resistance to Plutella xylostella through the accumulation of the homoterpene DMNT.

The lepidopteran crop pest Plutella xylostella causes severe constraints on Brassica cultivation. Here, we report a novel role for RPX1 (resistance to P. xylostella) in resistance to this pest in Arabidopsis thaliana. The rpx1-1 mutant repels P. xylostella larvae, and feeding on the rpx1-1 mutant severely damages the peritrophic matrix structure in the midgut of the larvae, thereby negatively affecting larval growth and pupation. This resistance results from the accumulation of defence compounds, including the homoterpene (3E)-4,8-dimethyl-1,3,7-nonatriene (DMNT), due to the upregulation of PENTACYCLIC TRITERPENE SYNTHASE 1 (PEN1), which encodes a key DMNT biosynthetic enzyme. P. xylostella infestation and wounding induce RPX1 protein degradation, which may confer a rapid response to insect infestation. RPX1 inactivation and PEN1 overexpression are not associated with negative trade-offs for plant growth but have much higher seed production than the wild-type in the presence of P. xylostella infestation. This study offers a new strategy for plant molecular breeding against P. xylostella.

Melatonin treatment delays postharvest senescence of broccoli with regulation of carotenoid metabolism.

The effect of melatonin treatment on the carotenoid metabolism in broccoli florets during storage was explored. The results indicated that 100 µmol/L of melatonin maintained the sensory quality of broccoli florets, which retarded the increase of the L* value and the decrease of the H value. Melatonin treatment increased the activities of tryptophan decarboxylase (TDC), tryptamine 5-hydroxylase (T5H), serotonin N-acetyltransferase (SNAT) and N-acetylserotonin methyltransferase (ASMT), leading to the enrichment of endogenous melatonin content in broccoli florets. Meanwhile, the treatment inhibited the concentrations of ß-carotene, ß-cryptoxanthin, zeaxanthin and lutein, which was beneficial in delaying the yellowing of broccoli. In addition, a series of carotenoid biosynthetic genes such as BoPSY, BoPDS, BoZDS, BoLCYß and BoZEP was also suppressed by melatonin. Further analysis revealed that the lower carotenoid content and the down-regulated BoNCED expression in treated broccoli resulted in less accumulation of abscisic acid precursors, inhibiting abscisic acid production during the yellowing process.

Rhizosphere bacteria regulated arsenic bioavailability and accumulation in the soil-Chinese cabbage system.

The accumulation of arsenic (As) in Chinese cabbage (Brassica rapa ssp. pekinensis) has recently been a source of concern for a potential risk to human health. It is unknown whether natural variations of As accumulation in different genotypes of Chinese cabbage are related to rhizobacterial characteristics. Experiments were conducted to investigate the mechanisms of rhizobacteria involving in As fates in a soil-Chinese cabbage system using various genotypes using high-throughput sequencing and quantitative PCR. There were significant differences in As accumulation in cabbage leaves between 32 genotypes, and genotypes of low-As-accumulation (LSA) and high-As-accumulation (HSA) were identified. The As concentrations in the shoots of LSA were 23.25 %, 24.19 %, 15.05 %, and 70.69 % lower than those of HSA in seedling stage (SS), rosette stage (RS), heading stage (HS), and mature stage (MS), respectively. Meanwhile, the relative abundances of phyla Patescibacteria (in RS), Acidobacteria and Rokubacteria (in HS) in the rhizosphere of LSA were 60.18 %, 28.19 %, and 45.38 % less than those of HSA, respectively. Additionally, both shoot-As and As translocation factor had significantly positive or negative correlations with the relative abundances of Rokubacteria or Actinobacteria. In LSA rhizosphere, the relative abundances of genera Flavobacterium (in SS), Ellin6055 and Sphingomonas (in HS) were 128.12 %, 83.69 % and 79.50 % higher than those of HSA, respectively. This demonstrated that rhizobacteria contribute to the accumulation and translocation of As in HSA and LSA. Furthermore, the gene copies of aioA and arsM in LSA rhizosphere were 25.54 % and 16.13 % higher than those of HSA, respectively, whereas the gene copies of arsC in LSA rhizosphere were 26.36 % less than those of HSA in MS, indicating that rhizobacteria are involved in As biotransformation in the soil. These results provide a comprehensive understanding of the relationship between characteristics of rhizobacterial communities and As variations in Chinese cabbage genotypes.

Conservation and Divergence of the Trihelix Genes in Brassica and Expression Profiles of BnaTH Genes in Brassica napus under Abiotic Stresses.

Trihelix (TH) proteins are a family of plant-specific transcription factors that play a role in light response and are extensively involved in plant growth and development, as well as in various stress responses. However, the function of TH genes in Brassica napus (B. napus) remains unclear, as does the evolution and differentiation pattern of TH genes in Brassica plants. Here, we identified a total of 455 TH genes in seven species, including six Brassica species and Arabidopsis, which were grouped into five clades, GT-1, GT-2, GTγ, SH4, and SIP1, each with 69, 142, 44, 55, and 145 members, respectively. The types and distributions of motifs of the TH proteins and the structures of the TH genes are conserved in the same subgroup, and some variations in certain amino acid residues occur in B. napus when inheriting motifs from Brassica rapa (B. rapa) and Brassica oleracea (B. oleracea). Collinearity analysis revealed that the massive expansion of TH genes in tetraploid species was attributed to the hetero-tetraploidization of diploid ancestors and gene duplication events within the tetraploid species. Comparative analysis of the membership numbers of five subgroups in different species revealed that the GT-2 and SIP1 genes underwent significant expansion during evolution, possibly to support the better adaptation of plants to their environments. The differential expression of the BnaTH genes under five stresses indicates that the BnaTH genes are involved in plant responses to stresses such as drought, cold, and heat. The presence of different stress-responsive cis-elements in the upstream promoter region of the genes indicated that BnaTH genes have the potential to cope with variable environments. Meanwhile, qRT-PCR analyses also confirmed that five TH genes respond to different abiotic stresses. Our results provide information and candidates for further studies on the role of TH genes in stress resistance of B. napus.

BocODD1 and BocODD2 Regulate the Biosynthesis of Progoitrin Glucosinolate in Chinese Kale.

Progoitrin (2-hydroxy-3-butenyl glucosinolate, PRO) is the main source of bitterness of Brassica plants. Research on the biosynthesis of PRO glucosinolate can aid the understanding of the nutritional value in Brassica plants. In this study, four ODD genes likely involved in PRO biosynthesis were cloned from Chinese kale. These four genes, designated as BocODD1-4, shared 75-82% similarities with the ODD sequence of Arabidopsis. The sequences of these four BocODDs were analyzed, and BocODD1 and BocODD2 were chosen for further study. The gene BocODD1,2 showed the highest expression levels in the roots, followed by the leaves, flowers, and stems, which is in accordance with the trend of the PRO content in the same tissues. Both the expression levels of BocODD1,2 and the content of PRO were significantly induced by high- and low-temperature treatments. The function of BocODDs involved in PRO biosynthesis was identified. Compared with the wild type, the content of PRO was increased twofold in the over-expressing BocODD1 or BocODD2 plants. Meanwhile, the content of PRO was decreased in the BocODD1 or BocODD2 RNAi lines more than twofold compared to the wildtype plants. These results suggested that BocODD1 and BocODD2 may play important roles in the biosynthesis of PRO glucosinolate in Chinese kale.

Biological Investigation and Chemical Study of Brassica villosa subsp. drepanensis (Brassicaeae) Leaves.

Brassica villosa subsp. drepanensis (Caruel) Raimondo & Mazzola, belonging to the Brassica oleracea complex, is a wild edible plant endemic to western Sicily and a relative of modern cultivated Brassica crops. In this study, the antioxidant properties, anti-inflammatory activities, enzymatic inhibition, and cytotoxicity in cancer cells of B. villosa subsp. drepanensis leaf ethanolic extract were analysed for the first time. In addition, its chemical profile was investigated partitioning the total 70% ethanol extract among ethyl acetate, n-butanol, and water to obtain three residues that were subjected to chromatographic separation. Two flavonol glycosides, a phenol glucoside, two amino acids, and purine/pyrimidine bases were obtained. The presence of the glucosinolate glucoiberin was detected in the water extract by UHPLC-MS analysis. The total polyphenol and flavonoid content of the 70% ethanol extract showed good antioxidant capacities and anti-inflammatory properties by reducing nitric oxide release and reactive oxygen species levels and increasing glutathione in lipopolysaccharide-stimulated RAW 264.7 cells. The extract inhibited the enzymatic activity of α-amylase, α-glucosidase, and, significantly, of lipase. The MTT assay showed that the extract did not affect the viability of normal HFF-1 and RAW 264.7 cells. Among the cancer cell lines tested, an antiproliferative action was only observed in CaCo-2. The cytotoxicity of the extract was further confirmed by LDH release assay and by the destabilization of the oxidative balance. Results confirmed the antioxidant properties of the crude extract responsible for the anti-inflammatory effect on healthy cells and cytotoxicity in cancer cells.