A 43 Bp-Deletion in the F3'H Gene Reducing Anthocyanins Is Responsible for Keeping Buds Green at Low Temperatures in Broccoli.

Most broccoli cultivars or accessions exhibit green buds under appropriate growth conditions, which turn purple at cold temperatures. However, certain cultivars consistently maintain green buds both during normal growth and at cold temperatures. In this study, we used BSA-seq (bulked segregation analysis-sequencing), along with fine mapping and transcriptome analysis to identify a candidate gene (flavonoid 3'-hydroxylase, F3'H) responsible for reducing anthocyanin accumulation in the mutant GS and HX-16 broccoli (Brassica oleracea L. var. italica), which could retain green buds even at low temperatures. A 43-bp deletion was detected in the coding sequence (CDS) of the F3'H gene in HX-16 and the mutant GS, which significantly decreased F3'H expression and the accumulation of cyanidin and delphinidin in the mutant GS. Furthermore, the expression of F3'H was upregulated at low temperatures in the wild line PS. Our results demonstrated the efficacy of utilizing the 43-bp InDel (Insertion-Deletion) in predicting whether buds in B. oleracea L. will turn purple or remain green at cold temperatures across forty-two germplasm materials. This study provides critical genetic and molecular insights for the molecular breeding of B. oleracea and sheds light on the molecular mechanisms underlying the effect of low temperatures on bud color in broccoli.

Comparative Transcriptome and Targeted Metabolome Profiling Unravel the Key Role of Phenylpropanoid and Glucosinolate Pathways in Defense against Alternaria brassicicola in Broccoli.

Alternaria brassicicola (Ab) can cause a major yield and quality-limiting disease of Brassica oleracea called black spot, and the genetic resources conferring complete resistance against Ab have not been identified to date. Here, comparative transcriptome and targeted metabolome analysis were performed utilizing a newly identified resistant (R) line and a broccoli susceptible (S) line at 6, 24, and 72 h post-inoculation (hpi). Kyoto encyclopedia of genes and genomes pathway enrichment and the weighted gene co-expression network analyses showed that the phenylpropanoid pathway regulates the resistance to Ab in broccoli. One metabolite, cinnamic acid, was significantly upregulated in the Ab_inoculated R line compared with the mock treatment but no significant difference in the S line, indicating that the cinnamic acid may cause the resistance difference between R and S lines. Our results also revealed that three indolic glucosinolates of I3G, 4MI3G, and 1MI3G were significantly increased in the Ab_inoculated R line compared with the mock treatment, and some related genes were differentially expressed between the R and S lines. These results provided new insights into the mechanism of Ab defense in B. oleracea and have laid a theoretical foundation for effectively utilizing resistant germplasm resources in broccoli breeding.

Molecular Markers for Detecting Inflorescence Size of Brassica oleracea L. Crops and B. oleracea Complex Species (n = 9) Useful for Breeding of Broccoli (B. oleracea var. italica) and Cauliflower (B. oleracea var. botrytis).

The gene flow from Brassica oleracea L. wild relatives to B. oleracea vegetable crops have occurred and continue to occur ordinarily in several Mediterranean countries, such as Sicily, representing an important hot spot of diversity for some of them, such as broccoli, cauliflower and kale. For detecting and for exploiting the forgotten alleles lost during the domestication processes of the B. oleracea crops, attention has been pointed to the individuation of specific markers for individuating genotypes characterized by hypertrophic inflorescence traits by the marker assisted selection (MAS) during the first plant growing phases after the crosses between broccoli (B. oleracea var. italica)/cauliflower (B. oleracea var. botrytis) with B. oleracea wild relatives (n = 9), reducing the cultivation and evaluation costs. The desired traits often found in several B. oleracea wild relatives are mainly addressed to improve the plant resistance to biotic and abiotic stresses and to increase the organoleptic, nutritive and nutraceutical traits of the products. One of the targeted traits for broccoli and cauliflower breeding is represented by the inflorescences size as is documented by the domestication processes of these two crops. Based on the previous results achieved, the numerical matrix, obtained utilizing five simple sequence repeats (SSRs), was analyzed to assess the relationship among the main inflorescence characteristics and the allelic variation of the SSRs loci analyzed (BoABI1, BoAP1, BoPLD1, BoTHL1 and PBCGSSRBo39), both for the Brassica oleracea and B. oleracea wild relatives (n = 9) accessions set. The main inflorescence morphometric characteristics, such as weight, height, diameter, shape, inflorescence curvature angle and its stem diameter, were registered before the flower anthesis. We analyzed the correlations among the allelic variation of the SSRs primers utilized and the inflorescence morphometric characteristics to individuate genomic regions stimulating the hypertrophy of the reproductive organ. The relationships found explain the diversity among B. oleracea crops and the B. oleracea complex species (n = 9) for the inflorescence size and structure. The individuated markers allow important time reduction during the breeding programs after crossing wild species for transferring useful biotic and abiotic resistances and organoleptic and nutraceutical traits to the B. oleracea crops by MAS.

Advances and Challenges for QTL Analysis and GWAS in the Plant-Breeding of High-Yielding: A Focus on Rapeseed.

Yield is one of the most important agronomic traits for the breeding of rapeseed (Brassica napus L), but its genetic dissection for the formation of high yield remains enigmatic, given the rapid population growth. In the present review, we review the discovery of major loci underlying important agronomic traits and the recent advancement in the selection of complex traits. Further, we discuss the benchmark summary of high-throughput techniques for the high-resolution genetic breeding of rapeseed. Biparental linkage analysis and association mapping have become powerful strategies to comprehend the genetic architecture of complex agronomic traits in crops. The generation of improved crop varieties, especially rapeseed, is greatly urged to enhance yield productivity. In this sense, the whole-genome sequencing of rapeseed has become achievable to clone and identify quantitative trait loci (QTLs). Moreover, the generation of high-throughput sequencing and genotyping techniques has significantly enhanced the precision of QTL mapping and genome-wide association study (GWAS) methodologies. Furthermore, this study demonstrates the first attempt to identify novel QTLs of yield-related traits, specifically focusing on ovule number per pod (ON). We also highlight the recent breakthrough concerning single-locus-GWAS (SL-GWAS) and multi-locus GWAS (ML-GWAS), which aim to enhance the potential and robust control of GWAS for improved complex traits.

Unraveling Sorghum Allelopathy in Agriculture: Concepts and Implications.

Allelopathy is an ecological phenomenon that involves the production and release of biomolecules from different crops, cultivated plants, and bacteria or fungi into the soil rhizosphere and impacts other organisms in the vicinity. Sorghum possesses vital allelopathic characteristics due to which it produces and releases different biomolecules from its root hairs, stems, and grains. Several studies have reported that sorghum acts as an allelopathic crop, decreasing the growth and eco-physiological attributes of surrounding plants and weeds growing simultaneously or subsequently in the field. Sorghum allelopathy has been exploited in the context of green manure, crop rotations, cover crops, and intercropping or mulching, whereas plant aqueous extracts or powder might be an alternate method of weed control. A diverse group of allelochemicals, including benzoic acid, p-hydroxybenzoic acid, vanillic acid, ferulic acid, chlorogenic acid, m-coumaric acid, p-coumaric acid, gallic acid, caffeic acid, p-hydroxibenzaldehyde, dhurrin, sorgoleone, m-hydroxybenzoic acid and protocatechuic acid, have been isolated and identified from different plant tissues of sorghum and root exudates. These allelochemicals, especially sorgoleone, have been investigated in terms of their mode(s) of action, specific activity and selectivity, release in the rhizosphere and uptake and translocation in sensitive species. The present review describes the importance of sorghum allelopathy as an ecological tool in managing weeds, highlighting the most recent advances in the allelochemicals present in sorghum, their modes of action, and their fate in the ecosystem. Further research should focus on the evaluation and selection of sorghum cultivars with high allelopathic potential, so that sorghum allelopathy can be better utilized for weed control and yield enhancement.

Effect of Drought Stress on Capsaicin and Antioxidant Contents in Pepper Genotypes at Reproductive Stage.

Pepper is one of the most important vegetables and spices in the world. Principal pungency is contributed by secondary metabolites called capsaicinoids, mainly synthesized in the placenta of pepper fruit. Various factors, including drought, limit pepper production. Flowering is one of the most sensitive stages affected by drought stress. The current study was conducted to determine the effect of drought on different pepper genotypes at the flowering and pod formation stages. Hot pepper (Pusajuala and Ghotki) and Bell pepper (Green Wonder and PPE-311) genotypes were subjected to drought (35% field capacity) at two different stages (flowering (DF) and pod formation (DP) stage). In comparison, control plants were maintained at 65% field capacity. The data regarding flowering survival rates, antioxidant protein activity, and proline content, were collected. Results indicated that parameters like flower survival percentage, number of fruits per plant, and fruit weight had significant differences among the genotypes in both treatments. A high proline level was observed in Green Wonder at the pod formation stage compared to other genotypes. Capsaicin contents of hot pepper genotypes were affected at the pod formation stage. Antioxidants like GPX were highly active (190 units) in Ghotki at pod formation. Bell pepper genotypes had a high APX activity, highly observed (100 units) in PPE-311 at pod formation, and significantly differ from hot pepper genotypes. In the catalase case, all the genotypes had the highest values in DP compared to control and DF, but Pusajuala (91 units) and Green Wonder (83 units) performed best compared to other genotypes. Overall, the results indicate that drought stress decreased reproductive growth parameters and pungency of pepper fruit as most of the plant energy was consumed in defense molecules (antioxidants). Therefore, water availability at the flowering and pod formation stage is critical to ensure good yield and pepper quality.

An Evaluation of Different Parameters to Screen Ornamental Shrubs for Salt Spray Tolerance.

In the context of seaside landscaping, one of the greatest challenges for practitioners and scientists is to select suitable species that are able to tolerate salt spray. This is the key aspect for a wide number of potentially suitable species. The objectives of this study were (1) to identify plant traits associated with species tolerance to salt stress and (2) to evaluate the responses of different shrub species to salt spray. For this purpose, a study was conducted to determine the effects of salt spray on twenty-four ornamental shrubs using rapid and easy-to-use methodology. The species were subjected twice a week to nebulization treatment with simulated seawater solution for 60 days. Every 20 days, net photosynthesis, stomatal conductance, transpiration rate, and chlorophyll a fluorescence were determined. Furthermore, dry biomass of the different organographic portions, leaf number, leaf area, Specific Leaf Area, chlorophyll content, and leaf damage were determined at the end of the experiment. The species exposed to seawater solution showed different physiological and morphological responses. Based on the above indices, these ornamental shrubs were screened and categorized as tolerant, moderately tolerant, or susceptible. The results suggest that Convolvulus, Ceratonia, and Ligustrum are more tolerant to salt spray than numerous other genotypes; L. langmaniae, Cascabela, and L. frutescens, conversely, are more sensitive. Among the plant traits, the morphological parameters thoroughly characterized the effects of the salt spray, but they were destructive, with the only exception being the leaf damage percentage. This last non-destructive parameter is interesting considering the aesthetic value that ornamental plants must have. The physiological parameters, and in particular photosynthesis activity, can instead be used as a non-destructive screening method to select species suitable for ornamental green spaces near the sea.

Phytochemical Characterization and In Vitro Antioxidant Properties of Four Brassica Wild Species from Italy.

In the present study, we evaluated for the first time the variability of antioxidant traits of four Brassica wild species: B. incana, B. macrocarpa, B. villosa, and B. rupestris. The content of the main water-soluble antioxidants (phenolics, ascorbic acid, and total biothiols) and the in vitro antioxidant potential (1,1-diphenyl-2-picrylhydrazil (DPPH) and superoxide anion scavenging capacity) were investigated. A total of 28 polyphenolic compounds were identified by LC/MS and quantitated by HPLC/DAD analysis. Kaempferol and quercetin derivatives were the most abundant phenolics compared to hydroxycinnamoyl gentiobiosides. In the ten populations, phenolics ranged from 163.9 to 533.9 mg/100 g dry weight (d.w.), ascorbic acid from 7.6 to 375.8 mg/100 g d.w., and total biothiols from 0.59 to 5.13 mg/100 g d.w. The different classes of phytochemicals were separated using solid-phase extraction at increasing methanol concentrations, and the antioxidant power of fractionated extracts was evaluated. The superoxide anion scavenging activity was significantly correlated to phenolics, particularly to flavonol derivatives, while DPPH was mainly related to ascorbic acid content. The present findings improve the knowledge of the phytochemical composition of Italian Brassica wild species by showing the great diversity of phytochemicals among populations and highlighting their importance as a valuable genetic resource for developing new cultivars with improved bioactive content.

Construction of a high-density genetic map and identification of loci controlling purple sepal trait of flower head in Brassica oleracea L. italica.

BACKGROUND: Some broccoli (Brassica oleracea L. italic) accessions have purple sepals and cold weather would deepen the purple color, while the sepals of other broccoli lines are always green even in cold winter. The related locus or gene is still unknown. In this study, a high-density genetic map was constructed based on specific locus amplified fragment (SLAF) sequencing in a doubled-haploid segregation population with 127 individuals. And mapping of the purple sepal trait in flower heads based on phenotypic data collected during three seasons was performed. RESULTS: A genetic map was constructed, which contained 6694 SLAF markers with an average sequencing depth of 81.37-fold in the maternal line, 84-fold in the paternal line, and 15.76-fold in each individual population studied. In all of the annual data recorded, three quantitative trait loci (QTLs) were identified that were all distributed within the linkage group (LG) 1. Among them, a major locus, qPH.C01-2, located at 36.393 cM LG1, was consistently detected in all analysis. Besides this locus, another two minor loci, qPH.C01-4 and qPH.C01-5, were identified near qPH.C01-2, based on the phenotypic data from spring of 2018. CONCLUSION: The purple sepal trait could be controlled by a major single locus and two minor loci. The genetic map and location of the purple sepal trait of flower heads provide an important foundation for mapping other compound traits and the identification of the genes related to purple sepal trait in broccoli.

Construction of a High-Density Genetic Map and Identification of Loci Related to Hollow Stem Trait in Broccoli (Brassic oleracea L. italica).

A high-quality genetic map is important for mapping of compound traits. In this study, a genetic map was constructed based on the reference genome TO1000 after specific locus amplified fragment (SLAF) sequencing in a double-haploid segregation population of broccoli, and loci controlling hollow stem trait were identified in the genetic map. The genetic map contains 4,787 SLAF markers, with a mean marker distance of 0.22 cM and the mean sequencing depths of 91.14-fold in the maternal line, 88.97-fold in the paternal line and 17.11-fold in each DH progeny. A locus controlling the hollow stem trait, QHS.C09-2, which could explain 14.1% of the phenotypic variation, was steadily detected on the linkage group nine in the indicated data of 3 years' trials and BLUE analysis. The genetic map could lay an important foundation for mapping of compound traits, and mapping of hollow stem trait would be basis to clone the genes related to hollow stems in broccoli.

An insight from tolerance to salinity stress in halophyte Portulaca oleracea L.: Physio-morphological, biochemical and molecular responses.

Salinity represents one of the environmental conditions with adverse effects on the productivity of most crops throughout the world. The response of plants to salt stress is of great interest for research to understand the mechanism involved in tolerance to salinity and highlight insights into the improvement of salt tolerance-crops of importance. In this study, the effect of salt stress was observed in wild and cultivated populations of P. oleracea originated from Tunisia and Italy. The results showed that at various concentrations of NaCl (0 mM, 50 mM, 100 mM and 150 mM), salinity has led to changes in growth parameters marked mainly by an increase in fresh and dry biomass. Beside, one of the salinity-induced side effects corresponds to the competition of Na+ and K+ ions for potassium root transporters. Our results suggested that purslane deployed an important element of tolerance such as the transporters ability to discriminate cations. In addition, the variation of PC5S gene expression tested by semi-quantitative RT-PCR revealed that proline synthesis is important in plants adaptation in saline conditions. A correlation between the gene expression varying by population and saline concentration and the level of proline assayed on the leaves of P. oleracea was highlighted.

The effect of the germination temperature on the phytochemical content of broccoli and rocket sprouts.

This study investigates the effect of different germination temperatures (10, 20 and 30 °C) on the phytochemical content as well as reducing and antioxidant capacity of broccoli and rocket sprouts. In both seeds and sprouts, the total glucosinolates and ascorbic acid contents did not differ between vegetables, while broccoli exhibited exceptionally higher polyphenols and greater reducing and antioxidant capacity compared to rocket. In both species, an increase in germination temperature positively affected the glucosinolate content. Ascorbic acid increased during germination without a difference among the three tested temperatures. The phenol content in broccoli sprouts increased when they were grown at 30 °C, but the amount decreased at the highest temperatures in rocket. The reducing and antioxidant capacities increased with germination, and higher indexes were detected at 10 °C, particularly in rocket. Different germination temperatures differentiate the health-promoting phytochemical content and antioxidant properties in broccoli and rocket sprouts.

Isatis canescens is a rich source of glucobrassicin and other health-promoting compounds.

BACKGROUND: Glucobrassicin (GBS), a glucosinolate contained in many brassica vegetables, is the precursor of chemopreventive compounds such as indole-3-carbinol. Large amounts of GBS would be needed to perform studies aimed at elucidating its role in the diet. This study was mainly undertaken to evaluate the flower buds of Isatis canescens as a source for GBS purification. In order to investigate the health-promoting potential of this species, glucosinolate, phenol and flavonoid content as well as the whole antioxidant capacity were also determined. Flower bud samples were collected in four localities around Mount Etna in Sicily, Italy, where I. canescens is widespread, as they are locally traditionally eaten. RESULTS: I. canescens flower buds displayed high GBS concentrations, up to 60 µmol g(-1) dry weight. The purification method consisted of two chromatographic steps, which made it possible to obtain GBS with a purity of 92-95%, with a yield of 21 g kg(-1) . The total glucosinolates, phenols, flavonoids and antioxidant activity were considerable, with the southern locality showing the highest concentrations for all the phytochemicals. CONCLUSION: I. canescens flower buds represent a naturally rich source of GBS, at a level suitable for its purification. Furthermore, flower bud consumption could provide an intake of health-promoting compounds, with possible antioxidant and chemopreventive properties.

Genetic diversity and population structure of leafy kale and Brassica rupestris Raf. in south Italy.

Local varieties of leafy kales (Brassica oleracea L.) are grown in home gardens in Calabria and Sicily for self-consumption, in the same area where the wild relative Brassica rupestris Raf. also grows. With the use of AFLP markers, comparisons were made of the genetic diversity and population structure of ten wild and 22 cultivated populations, as well as of a hybrid population and of four commercial cultivars of different B. oleracea crops. The level of genetic diversity was higher in leafy kales than in wild populations and this diversity was mainly distributed within populations. Wild populations remained distinct from cultivated material. Additionally, most wild populations were distinctively isolated from each other. On the other hand, it was not possible to molecularly distinguish even geographically distant leafy kale populations from each other or from different B. oleracea crops. It was possible to detect inter-crossing between leafy kales and B. rupestris. Findings from this study illustrate the existing level of genetic diversity in the B. oleracea gene pool. Individual populations (either wild or leafy kales) with higher levels of genetic diversity have been identified and suggestions are given for an informed conservation strategy. Domestication hypotheses are also discussed.

Study and characterization of polyphenol oxidase from eggplant (Solanum melongena L.).

In this study the catecholase and cresolase activities of eggplant polyphenol oxidase (PPO) were investigated. Enzyme activity was determined by measuring the increase in absorbance using catechol as substrate and 3-methyl-2-benzothiazolinone hydrazone (MBTH) as coupled reagent. The effects of substrate specificity, heat inactivation, temperature, pH, and inhibitors were investigated to understand the enzymatic alteration of ready-to-eat preparations. Browning of vegetables was determined through a colorimeter. Decrease of lightness (L*) and increase of color difference values (ΔE*) were correlated with tissue browning. Antibrowning agents were tested on PPO under the same conditions. The enzyme activity was strongly inhibited by 0.4 M citric acid. Under natural pH conditions, the enzyme was also inhibited by tartaric acid and acetic acid. All of the results were used to understand the best conditions for food transformation (ready-to-eat and grilled eggplant slices).

Characterization of a tomato polyphenol oxidase and its role in browning and lycopene content.

Polyphenol oxidase (PPO) was extracted from five Sicilian varieties of tomato fruit [Pizzutello, Naomi (Hazera), F1 PS212 (Peto seed), Rosa Maletto, and PO228] and assayed with a method using 3-methylbenzothyazolinone hydrazone (MBTH) as chromophore coupling agent. 3,4-Dihydroxyphenylacetic acid was chosen for tomato PPO activity determination. The tomato PPO had maximum activity at pH 4.8. The pH of juice in ripe fruits is between 4.1 and 4.4, a range in which PPO relative activity is between 74 and 87%. The optimum temperature of activity for tomato PPO was 40 degrees C; the enzyme showed a good relative activity (55% of the maximum) at cold-storage temperature (4 degrees C). PPO retained 82% relative activity at an NaCl concentration of 0.1 M; at higher concentrations the PPO became gradually inactivated. The commercial variety Naomi is more susceptible to enzymatic browning than the local varieties Pizzutello, Rosa Maletto and PO228, due to higher PPO activity levels. This result confirms the suitability of these local tomato varieties to national markets. Results from storage tests seem to relate PPO activity with color changes associated with browning and lycopene degradation, because lycopene is an antioxidant agent that reconstitutes the polyphenols oxidized by the action of PPO.

Survey of aliphatic glucosinolates in Sicilian wild and cultivated Brassicaceae.

In the frame of the activities carried out to exploit Sicilian local cultivars of brassicas, we focused our attention on some of the potential health compounds of various local cruciferous crops. These compounds are of interest to improve the quality of the produce with the aim to develop new cultivars capable of providing functional foods able to prevent disease. In this context, we surveyed for the presence of specific glucosinolates in local cultivars of broccoli, cauliflower, kale, and in some wild species widespread in Sicily, using as control various commercial cultivars. Glucosinolate composition varied extensively among species and crops of the same species, such as cauliflower, broccoli and kale. Cultivar variation for glucosinolate profile was also observed for some crops. For example, Sicilian cultivars of cauliflower possessing colored curds displayed a high content of glucosinolates, glucoraphanin in particular, compared to white curd commercial cultivars. Also some wild species had a high content of other glucosinolates.