Insights into chloroplast genome structure and phylogenetic relationships within the Sesamum species complex (Pedaliaceae).

Background: In the Sesamum species complex, the lack of wild species genomic resources hinders the evolutionary comprehension of phylogenetic relationships. Results: In the present study, we generated complete chloroplast genomes of six wild relatives (Sesamum alatum, Sesamum angolense, Sesamum pedaloides, Ceratotheca sesamoides (syn. Sesamum sesamoides), Ceratotheca triloba (syn. Sesamum trilobum), and Sesamum radiatum) and a Korean cultivar, Sesamum indicum cv. Goenbaek. A typical quadripartite chloroplast structure, including two inverted repeats (IR), a large single copy (LSC), and a small single copy (SSC), was observed. A total of 114 unique genes encompassing 80 coding genes, four ribosomal RNAs, and 30 transfer RNAs were counted. The chloroplast genomes (152, 863-153, 338 bp) exhibited the IR contraction/expansion phenomenon and were quite conserved in both coding and non-coding regions. However, high values of the nucleotide diversity index were found in several genes, including ndhA, ndhE, ndhF, ycf1, and psaC-ndhD. Concordant tree topologies suggest ndhF as a useful marker for taxon discrimination. The phylogenetic inference and time divergence dating indicate that S. radiatum (2n = 64) occurred concomitantly with the sister species C. sesamoides (2n = 32) approximately 0.05 million years ago (Mya). In addition, S. alatum was clearly discriminated by forming a single clade, showing its long genetic distance and potential early speciation event in regards to the others. Conclusion: Altogether, we propose to rename C. sesamoides and C. triloba as S. sesamoides and S. trilobum, respectively, as suggested previously based on the morphological description. This study provides the first insight into the phylogenetic relationships among the cultivated and wild African native relatives. The chloroplast genome data lay a foundation for speciation genomics in the Sesamum species complex.

Characteristics and Fitness Analysis through Interspecific Hybrid Progenies of Transgenic Brassica napus and B. rapa L. ssp.

Interspecific hybridization between transgenic crops and their wild relatives is a major concern for transgene dispersal in the environment. Under controlled conditions, artificial hand pollination experiments were performed in order to assess the hybridization potential and the fitness of interspecific hybrids between Brassica rapa and genetically modified (GM) Brassica napus. Initially, six subspecies of B. rapa were hybridized with GM B. napus through hand pollination. In the resulting F1 hybrids, the combination of B. rapa ssp. narinosa (♀) × GM B. napus (♂) had the highest crossability index (16.9 ± 2.6). However, the F1 selfing progenies of B. rapa ssp. rapa (♀) × GM B. napus were found to be more effective in producing viable future generations with the highest crossability index (1.6 ± 0.69) compared to other subspecies. Consequently, they were used for the generation of F2 and F3 progenies. The 18 different morphological characteristics among the parental cross-combinations and F1 hybrid progenies were measured and visualized through hierarchical clustering. Different generations were found to be grouped based on their different morphological characteristics. The chromosome numbers among the interspecific hybrids ranged from 2n = 29 to 2n = 40. Furthermore, the SSR markers revealed the presence of genomic portions in the hybrids in comparison with their parental lines. There is a high possibility of transgene flow between GM B. napus and B. rapa. The study concluded that the interspecific hybrids between B. napus and B. rapa can be viable and can actively hybridize up to F3 generations and more. This suggests that the GM B. napus can disperse the transgene into B. rapa, and that it can pass through for several generations by hand pollination in a greenhouse environment.

Interspecific Hybridization of Transgenic Brassica napus and Brassica rapa-An Overview.

In nature, interspecific hybridization occurs frequently and can contribute to the production of new species or the introgression of beneficial adaptive features between species. It has great potential in agricultural systems to boost the process of targeted crop improvement. In the advent of genetically modified (GM) crops, it has a disadvantage that it involves the transgene escaping to unintended plants, which could result in non-specific weedy crops. Several crop species in the Brassica genus have close kinship: canola (Brassica napus) is an ancestral hybrid of B. rapa and B. oleracea and mustard species such as B. juncea, B. carinata, and B. nigra share common genomes. Hence, intraspecific hybridization among the Brassica species is most common, especially between B. napus and B. rapa. In general, interspecific hybrids cause numerous genetic and phenotypic changes in the parental lines. Consequently, their fitness and reproductive ability are also highly varied. In this review, we discuss the interspecific hybridization and reciprocal hybridization studies of B. napus and B. rapa and their potential in the controlled environment. Further, we address the fate of transgenes (herbicide resistance) and their ability to transfer to their progenies or generations. This could help us to understand the environmental influence of interspecific hybrids and how to effectively manage their transgene escape in the future.

Jasmonates in plant growth and development and elicitation of secondary metabolites: An updated overview.

Secondary metabolites are incontestably key specialized molecules with proven health-promoting effects on human beings. Naturally synthesized secondary metabolites are considered an important source of pharmaceuticals, food additives, cosmetics, flavors, etc., Therefore, enhancing the biosynthesis of these relevant metabolites by maintaining natural authenticity is getting more attention. The application of exogenous jasmonates (JAs) is well recognized for its ability to trigger plant growth and development. JAs have a large spectrum of action that covers seed germination, hypocotyl growth regulation, root elongation, petal expansion, and apical hook growth. This hormone is considered as one of the key regulators of the plant's growth and development when the plant is under biotic or abiotic stress. The JAs regulate signal transduction through cross-talking with other genes in plants and thereby deploy an appropriate metabolism in the normal or stressed conditions. It has also been found to be an effective chemical elicitor for the synthesis of naturally occurring secondary metabolites. This review discusses the significance of JAs in the growth and development of plants and the successful outcomes of jasmonate-driven elicitation of secondary metabolites including flavonoids, anthraquinones, anthocyanin, xanthonoid, and more from various plant species. However, as the enhancement of these metabolites is essentially measured via in vitro cell culture or foliar spray, the large-scale production is significantly limited. Recent advancements in the plant cell culture technology lay the possibilities for the large-scale manufacturing of plant-derived secondary metabolites. With the insights about the genetic background of the metabolite biosynthetic pathway, synthetic biology also appears to be a potential avenue for accelerating their production. This review, therefore, also discussed the potential manoeuvres that can be deployed to synthesis plant secondary metabolites at the large-scale using plant cell, tissue, and organ cultures.

Molecular characterisation and expression profiling of calcineurin B-like (CBL) genes in Chinese cabbage under abiotic stresses.

Calcium signals act as a second messenger in plant responses to various abiotic stresses, which regulate a range of physiological processes. Calcium-binding proteins, like calcineurin B-like (CBL) proteins, belong to a unique group of calcium sensors that play a role in calcium signalling. However, their identities and functions are unknown in Chinese cabbage. In this study, 17 CBL genes were identified from the Brassica rapa L. (Chinese cabbage) database and Br135K microarray datasets. They were used to construct a phylogenetic tree with known CBL proteins of other species. Analysis of genomic distribution and evolution revealed different gene duplication in Chinese cabbage compared to Arabidopsis. The microarray expression analysis showed differential expression of BrCBL genes at various temperatures. Organ-specific expression was observed by RT-PCR, and qRT-PCR analyses revealed responsiveness of BrCBL genes to cold, drought and salt stresses. Our findings confirm that CBL genes are involved in calcium signalling and regulate responses to environmental stimuli, suggesting this family gene have crucial role to play in plant responses to abiotic stresses. The results facilitate selection of candidate genes for further functional characterisation. In addition, abiotic stress-responsive genes reported in this study might be exploited for marker-aided backcrossing of Chinese cabbage.

De Novo Assembly and Transcriptome Analysis of Bulb Onion (Allium cepa L.) during Cold Acclimation Using Contrasting Genotypes.

Bulb onion (Allium cepa) is the second most widely cultivated and consumed vegetable crop in the world. During winter, cold injury can limit the production of bulb onion. Genomic resources available for bulb onion are still very limited. To date, no studies on heritably durable cold and freezing tolerance have been carried out in bulb onion genotypes. We applied high-throughput sequencing technology to cold (2°C), freezing (-5 and -15°C), and control (25°C)-treated samples of cold tolerant (CT) and cold susceptible (CS) genotypes of A. cepa lines. A total of 452 million paired-end reads were de novo assembled into 54,047 genes with an average length of 1,331 bp. Based on similarity searches, these genes were aligned with entries in the public non-redundant (nr) database, as well as KEGG and COG database. Differentially expressed genes (DEGs) were identified using log10 values with the FPKM method. Among 5,167DEGs, 491 genes were differentially expressed at freezing temperature compared to the control temperature in both CT and CS libraries. The DEG results were validated with qRT-PCR. We performed GO and KEGG pathway enrichment analyses of all DEGs and iPath interactive analysis found 31 pathways including those related to metabolism of carbohydrate, nucleotide, energy, cofactors and vitamins, other amino acids and xenobiotics biodegradation. Furthermore, a large number of molecular markers were identified from the assembled genes, including simple sequence repeats (SSRs) 4,437 and SNP substitutions of transition and transversion types of CT and CS. Our study is the first to provide a transcriptome sequence resource for Allium spp. with regard to cold and freezing stress. We identified a large set of genes and determined their DEG profiles under cold and freezing conditions using two different genotypes. These data represent a valuable resource for genetic and genomic studies of Allium spp.

Glutathione Transferases Superfamily: Cold-Inducible Expression of Distinct GST Genes in Brassica oleracea.

Plants, as sessile organisms, can suffer serious growth and developmental consequences under cold stress conditions. Glutathione transferases (GSTs, EC 2.5.1.18) are ubiquitous and multifunctional conjugating proteins, which play a major role in stress responses by preventing oxidative damage by reactive oxygen species (ROS). Currently, understanding of their function(s) during different biochemical and signaling pathways under cold stress condition remain unclear. In this study, using combined computational strategy, we identified 65 Brassica oleracea glutathione transferases (BoGST) and characterized them based on evolutionary analysis into 11 classes. Inter-species and intra-species duplication was evident between BoGSTs and Arabidopsis GSTs. Based on localization analyses, we propose possible pathways in which GST genes are involved during cold stress. Further, expression analysis of the predicted putative functions for GST genes were investigated in two cold contrasting genotypes (cold tolerance and susceptible) under cold condition, most of these genes were highly expressed at 6 h and 1 h in the cold tolerant (CT) and cold susceptible (CS) lines, respectively. Overall, BoGSTU19, BoGSTU24, BoGSTF10 are candidate genes highly expressed in B. oleracea. Further investigation of GST superfamily in B. oleracea will aid in understanding complex mechanism underlying cold tolerance in plants.

Whole Genome Re-Sequencing and Characterization of Powdery Mildew Disease-Associated Allelic Variation in Melon.

Powdery mildew is one of the most common fungal diseases in the world. This disease frequently affects melon (Cucumis melo L.) and other Cucurbitaceous family crops in both open field and greenhouse cultivation. One of the goals of genomics is to identify the polymorphic loci responsible for variation in phenotypic traits. In this study, powdery mildew disease assessment scores were calculated for four melon accessions, 'SCNU1154', 'Edisto47', 'MR-1', and 'PMR5'. To investigate the genetic variation of these accessions, whole genome re-sequencing using the Illumina HiSeq 2000 platform was performed. A total of 754,759,704 quality-filtered reads were generated, with an average of 82.64% coverage relative to the reference genome. Comparisons of the sequences for the melon accessions revealed around 7.4 million single nucleotide polymorphisms (SNPs), 1.9 million InDels, and 182,398 putative structural variations (SVs). Functional enrichment analysis of detected variations classified them into biological process, cellular component and molecular function categories. Further, a disease-associated QTL map was constructed for 390 SNPs and 45 InDels identified as related to defense-response genes. Among them 112 SNPs and 12 InDels were observed in powdery mildew responsive chromosomes. Accordingly, this whole genome re-sequencing study identified SNPs and InDels associated with defense genes that will serve as candidate polymorphisms in the search for sources of resistance against powdery mildew disease and could accelerate marker-assisted breeding in melon.

Characterization and abiotic stress-responsive expression analysis of SGT1 genes in Brassica oleracea.

SGT1 genes are involved in enhancing plant responses to various biotic and abiotic stresses. Brassica oleracea is known to contain two types of SGT1 genes, namely suppressor of G2 allele of SKP1 and suppressor of GCR2. In this study, through systematic analysis, four putative SGT1 genes were identified and characterized in B. oleracea. In phylogenetic analysis, the genes clearly formed separate groups, namely BolSGT1a, BolSGT1b (both suppressor of G2 allele of SKP1 types), and BolSGT1 (suppressor of GCR2). Functional domain analysis and organ-specific expression patterns suggested possible roles for BolSGT1 genes during stress conditions. BolSGT1 genes showed significant changes in expression in response to heat, cold, drought, salt, or ABA treatment. Interaction network analysis supported the expression analysis, and showed that the BolSGT1a and BolSGT1b genes are strongly associated with co-regulators during stress conditions. However, the BolSGT1 gene did not show any strong association. Hence, BolSGT1 might be a stress resistance-related gene that functions without a co-regulator. Our results show that BolSGT1 genes are potential target genes to improve B. oleracea resistance to abiotic stresses such as heat, cold, and salt.

Molecular and Functional Characterization of FLOWERING LOCUS T Homologs in Allium cepa.

Onion bulbing is an important agricultural trait affecting economic value and is regulated by flowering-related genes. FLOWERING LOCUS T (FT)-like gene function is crucial for the initiation of flowering in various plant species and also in asexual reproduction in tuber plants. By employing various computational analysis using RNA-Seq data, we identified eight FT-like genes (AcFT) encoding PEBP (phosphatidylethanolamine-binding protein) domains in Allium cepa. Sequence and phylogenetic analyses of FT-like proteins revealed six proteins that were identical to previously reported AcFT1-6 proteins, as well as one (AcFT7) with a highly conserved region shared with AcFT6 and another (comp106231) with low similarity to MFT protein, but containing a PEBP domain. Homology modelling of AcFT7 proteins showed similar structures and conservation of amino acids crucial for function in AtFT (Arabidopsis) and Hd3a (rice), with variation in the C-terminal region. Further, we analyzed AcFT expression patterns in different transitional stages, as well as under SD (short-day), LD (long-day), and drought treatment in two contrasting genotypic lines EM (early maturation, 36101) and LM (late maturation, 36122). The FT transcript levels were greatly affected by various environmental factors such as photoperiod, temperature and drought. Our results suggest that AcFT7 is a member of the FT-like genes in Allium cepa and may be involved in regulation of onion bulbing, similar to other FT genes. In addition, AcFT4 and AcFT7 could be involved in establishing the difference in timing of bulb maturity between the two contrasting onion lines.

Molecular Modeling of Myrosinase from Brassica oleracea: A Structural Investigation of Sinigrin Interaction.

Myrosinase, which is present in cruciferous plant species, plays an important role in the hydrolysis of glycosides such as glucosinolates and is involved in plant defense. Brassicaceae myrosinases are diverse although they share common ancestry, and structural knowledge about myrosinases from cabbage (Brassica oleracea) was needed. To address this, we constructed a three-dimensional model structure of myrosinase based on Sinapis alba structures using Iterative Threading ASSEmbly Refinement server (I-TASSER) webserver, and refined model coordinates were evaluated with ProQ and Verify3D. The resulting model was predicted with ß/α fold, ten conserved N-glycosylation sites, and three disulfide bridges. In addition, this model shared features with the known Sinapis alba myrosinase structure. To obtain a better understanding of myrosinase-sinigrin interaction, the refined model was docked using Autodock Vina with crucial key amino acids. The key nucleophile residues GLN207 and GLU427 were found to interact with sinigrin to form a hydrogen bond. Further, 20-ns molecular dynamics simulation was performed to examine myrosinase-sinigrin complex stability, revealing that residue GLU207 maintained its hydrogen bond stability throughout the entire simulation and structural orientation was similar to that of the docked state. This conceptual model should be useful for understanding the structural features of myrosinase and their binding orientation with sinigrin.

Genome-wide transcriptome analysis of two contrasting Brassica rapa doubled haploid lines under cold-stresses using Br135K oligomeric chip.

Genome wide transcription analysis in response to stresses is important to provide a basis of effective engineering strategies to improve stress tolerance in crop plants. We assembled a Brassica rapa oligomeric microarray (Br135K microarray) using sequence information from 41,173 unigenes and analyzed the transcription profiles of two contrasting doubled haploid (DH) lines, Chiifu and Kenshin, under cold-treatments. The two DH lines showed great differences in electrolyte leakage below -4°C, but similar patterns from 4°C to -2°C. Cold-treatments induced 885 and 858 genes in Chiifu and Kenshin, respectively. Overall, 134, and 56 genes showed an intrinsic difference in expression in Chiifu and Kenshin, respectively. Among 5,349 genes that showed no hit found (NHF) in public databases, 61 and 24 were specifically expressed in Chiifu and Kenshin, respectively. Many transcription factor genes (TFs) also showed various characteristics of expression. BrMYB12, BrMYBL2, BrbHLHs, BrbHLH038, a C2H2, a WRKY, BrDREB19 and a integrase-type TF were induced in a Chiifu-specific fashion, while a bHLH (Bra001826/AT3G21330), bHLH, cycling Dof factor and two Dof type TFs were Kenshin specific. Similar to previous studies, a large number of genes were differently induced or regulated among the two genotypes, but many genes, including NHFs, were specifically or intrinsically expressed with genotype specificity. Expression patterns of known-cold responsive genes in plants resulted in discrepancy to membrane leakage in the two DH lines, indicating that timing of gene expression is more important to conferring freezing tolerance rather than expression levels. Otherwise, the tolerance will be related to the levels of transcripts before cold-treatment or regulated by other mechanisms. Overall, these results indicate common signaling pathways and various transcriptional regulatory mechanisms are working together during cold-treatment of B. rapa. Our newly developed Br135K oligomeric microarray will be useful for transcriptome profiling, and will deliver valuable insight into cold stresses in B. rapa.

Genome-wide analysis of the distribution of AP2/ERF transcription factors reveals duplication and CBFs genes elucidate their potential function in Brassica oleracea.

BACKGROUND: Cabbage (Brassica oleracea) is one of the most important leaf vegetables grown worldwide. The entire cabbage genome sequence and more than fifty thousand proteins have been obtained to date. However a high degree of sequence similarity and conserved genome structure remain between cabbage and Arabidopsis; therefore, Arabidopsis is a viable reference species for comparative genomics studies. Transcription factors (TFs) are important regulators involved in plant development and physiological processes and the AP2/ERF protein family contains transcriptional factors that play a crucial role in plant growth and development, as well as response to biotic and abiotic stress conditions in plants. However, no detailed expression profile of AP2/ERF-like genes is available for B. oleracea. RESULTS: In the present study, 226 AP2/ERF TFs were identified from B. oleracea based on the available genome sequence. Based on sequence similarity, the AP2/ERF superfamily was classified into five groups (DREB, ERF, AP2, RAV and Soloist) and 15 subgroups. The identification, classification, phylogenetic construction, conserved motifs, chromosome distribution, functional annotation, expression patterns and interaction network were then predicted and analyzed. AP2/ERF transcription factor expression levels exhibited differences in response to varying abiotic stresses based on expressed sequence tags (ESTs). BoCBF1a, 1b, 2, 3 and 4, which were highly conserved in Arabidopsis and B. rapa CBF/DREB genes families were well characterized. Expression analysis enabled elucidation of the molecular and genetic level expression patterns of cold tolerance (CT) and susceptible lines (CS) of cabbage and indicated that all BoCBF genes responded to abiotic stresses. CONCLUSIONS: Comprehensive analysis of the physiological functions and biological roles of AP2/ERF superfamily genes and BoCBF family genes in B. oleracea is required to fully elucidate AP2/ERF, which will provide rich resources and opportunities to understand abiotic stress tolerance in crops.