Precise Determination of Selenium Forms and Contents in Selenium-Enriched Rapeseed Seedlings and Flowering Stalks by HPLC-ICP-MS.

Rapeseed (Brassica napus L.) has the ability of selenium (Se) enrichment. Identification of selenides in Se-rich rapeseed products will promote the development and utilization of high value. By optimizing the Se species extraction process (protease type, extraction reagent, enzyme sample ratio, extraction time, etc.) and chromatographic column, an efficient, stable, and accurate method was established for the identification of Se species and content in rapeseed seedlings and flowering stalks, which were cultured by inorganic Se hydroponics. Five Se compounds, including selenocystine (SeCys2), methylselenocysteine (MeSeCys), selenomethionine (SeMet), selenite (SeIV), and selenate (SeVI) were qualitatively and quantitatively identified. Organoselenium was absolutely dominant in both seedlings and flowering stalks among the detected rapeseed varieties, with 64.18-90.20% and 94.38-98.47%, respectively. Further, MeSeCys, a highly active selenide, predominated in rapeseed flowering stalks with a proportion of 56.36-72.93% and a content of 1707.3-5030.3 µg/kg. This study provides a new source of MeSeCys supplementation for human Se fortification.

An integrated QTL mapping and transcriptome sequencing provides further molecular insights and candidate genes for stem strength in rapeseed (Brassica napus L.).

KEY MESSAGE: We detected the major QTL- qSR.A07, which regulated stem strength and was fine-mapped to 490 kb. BnaA07G0302800ZS and BnaA07G0305700ZS as the candidate functional genes were identified at qSR.A07 locus. The stem's mechanical properties reflect its ability to resist lodging. In rapeseed (Brassica napus L.), although stem lodging negatively affects yield and generates harvesting difficulties, the molecular regulation of stem strength remains elusive. Hence, this study aimed to unravel the main loci and molecular mechanisms governing rapeseed stem strength. A mapping population consisting of 267 RILs (recombinant inbred lines) was developed from the crossed between ZS11 (high stem strength) and 4D122 (low stem strength), and two mechanical properties of stems including stem breaking strength and stem rind penetrometer resistance were phenotyped in four different environments. Four pleiotropic QTLs that were stable in at least two environments were detected. qSR.A07, the major one, was fine-mapped to a 490 kb interval between markers SA7-2711 and SA7-2760 on chromosome 7. It displayed epistatic interaction with qRPR.A09-2. Comparative transcriptome sequencing and analysis unveiled methionine/S-adenosylmethionine cycle (Met/SAM cycle), cytoskeleton organization, sulfur metabolism and phenylpropanoid biosynthesis as the main pathways associated with high stem strength. Further, we identified two candidate genes, BnaA07G0302800ZS and BnaA07G0305700ZS, at qSR.A07 locus. Gene sequence alignment identified a number of InDels, SNPs and amino acid variants in sequences of these genes between ZS11 and 4D122. Finally, based on these genetic variants, we developed three SNP markers of these genes to facilitate future genetic selection and functional studies. These findings offer important genetic resources for the molecular-assisted breeding of novel rapeseed stem lodging-resistant varieties.

Integrating genome-wide association study with transcriptomic data to predict candidate genes influencing Brassica napus root and biomass-related traits under low phosphorus conditions.

BACKGROUND: Rapeseed (Brassica napus L.) is an essential source of edible oil and livestock feed, as well as a promising source of biofuel. Breeding crops with an ideal root system architecture (RSA) for high phosphorus use efficiency (PUE) is an effective way to reduce the use of phosphate fertilizers. However, the genetic mechanisms that underpin PUE in rapeseed remain elusive. To address this, we conducted a genome-wide association study (GWAS) in 327 rapeseed accessions to elucidate the genetic variability of 13 root and biomass traits under low phosphorus (LP; 0.01 mM P +). Furthermore, RNA-sequencing was performed in root among high/low phosphorus efficient groups (HP1/LP1) and high/low phosphorus stress tolerance groups (HP2/LP2) at two-time points under control and P-stress conditions. RESULTS: Significant variations were observed in all measured traits, with heritabilities ranging from 0.47 to 0.72, and significant correlations were found between most of the traits. There were 39 significant trait-SNP associations and 31 suggestive associations, which integrated into 11 valid quantitative trait loci (QTL) clusters, explaining 4.24-24.43% of the phenotypic variance observed. In total, RNA-seq identified 692, 1076, 648, and 934 differentially expressed genes (DEGs) specific to HP1/LP1 and HP2/LP2 under P-stress and control conditions, respectively, while 761 and 860 DEGs common for HP1/LP1 and HP2/LP2 under both conditions. An integrated approach of GWAS, weighted co-expression network, and differential expression analysis identified 12 genes associated with root growth and development under LP stress. In this study, six genes (BnaA04g23490D, BnaA09g08440D, BnaA09g04320D, BnaA09g04350D, BnaA09g04930D, BnaA09g09290D) that showed differential expression were identified as promising candidate genes for the target traits. CONCLUSION: 11 QTL clusters and 12 candidate genes associated with root and development under LP stress were identified in this study. Our study's phenotypic and genetic information may be exploited for genetic improvement of root traits to increase PUE in rapeseed.

Research Progress of Selenium-Enriched Foods.

Selenium is an essential micronutrient that plays a crucial role in maintaining human health. Selenium deficiency is seriously associated with various diseases such as Keshan disease, Kashin-Beck disease, cataracts, and others. Conversely, selenium supplementation has been found to have multiple effects, including antioxidant, anti-inflammatory, and anticancer functions. Compared with inorganic selenium, organic selenium exhibits higher bioactivities and a wider range of safe concentrations. Consequently, there has been a significant development of selenium-enriched foods which contain large amounts of organic selenium in order to improve human health. This review summarizes the physiological role and metabolism of selenium, the development of selenium-enriched foods, the physiological functions of selenium-enriched foods, and provides an analysis of total selenium and its species in selenium-enriched foods, with a view to laying the foundation for selenium-enriched food development.

Transcriptome analysis reveals key regulatory genes for root growth related to potassium utilization efficiency in rapeseed (Brassica napus L.).

Root system architecture (RSA) is the primary predictor of nutrient intake and significantly influences potassium utilization efficiency (KUE). Uncertainty persists regarding the genetic factors governing root growth in rapeseed. The root transcriptome analysis reveals the genetic basis driving crop root growth. In this study, RNA-seq was used to profile the overall transcriptome in the root tissue of 20 Brassica napus accessions with high and low KUE. 71,437 genes in the roots displayed variable expression profiles between the two contrasting genotype groups. The 212 genes that had varied expression levels between the high and low KUE lines were found using a pairwise comparison approach. The Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) functional classification analysis revealed that the DEGs implicated in hormone and signaling pathways, as well as glucose, lipid, and amino acid metabolism, were all differently regulated in the rapeseed root system. Additionally, we discovered 33 transcription factors (TFs) that control root development were differentially expressed. By combining differential expression analysis, weighted gene co-expression network analysis (WGCNA), and recent genome-wide association study (GWAS) results, four candidate genes were identified as essential hub genes. These potential genes were located fewer than 100 kb from the peak SNPs of QTL clusters, and it was hypothesized that they regulated the formation of the root system. Three of the four hub genes' homologs-BnaC04G0560400ZS, BnaC04G0560400ZS, and BnaA03G0073500ZS-have been shown to control root development in earlier research. The information produced by our transcriptome profiling could be useful in revealing the molecular processes involved in the growth of rapeseed roots in response to KUE.

Integrating biochemical and anatomical characterizations with transcriptome analysis to dissect superior stem strength of ZS11 (Brassica napus).

Stem lodging resistance is a serious problem impairing crop yield and quality. ZS11 is an adaptable and stable yielding rapeseed variety with excellent resistance to lodging. However, the mechanism regulating lodging resistance in ZS11 remains unclear. Here, we observed that high stem mechanical strength is the main factor determining the superior lodging resistance of ZS11 through a comparative biology study. Compared with 4D122, ZS11 has higher rind penetrometer resistance (RPR) and stem breaking strength (SBS) at flowering and silique stages. Anatomical analysis shows that ZS11 exhibits thicker xylem layers and denser interfascicular fibrocytes. Analysis of cell wall components suggests that ZS11 possessed more lignin and cellulose during stem secondary development. By comparative transcriptome analysis, we reveal a relatively higher expression of genes required for S-adenosylmethionine (SAM) synthesis, and several key genes (4-COUMATATE-CoA LIGASE, CINNAMOYL-CoA REDUCTASE, CAFFEATE O-METHYLTRANSFERASE, PEROXIDASE) involved in lignin synthesis pathway in ZS11, which support an enhanced lignin biosynthesis ability in the ZS11 stem. Moreover, the difference in cellulose may relate to the significant enrichment of DEGs associated with microtubule-related process and cytoskeleton organization at the flowering stage. Protein interaction network analysis indicate that the preferential expression of several genes, such as LONESOME HIGHWAY (LHW), DNA BINDING WITH ONE FINGERS (DOFs), WUSCHEL HOMEOBOX RELATED 4 (WOX4), are related to vascular development and contribute to denser and thicker lignified cell layers in ZS11. Taken together, our results provide insights into the physiological and molecular regulatory basis for the formation of stem lodging resistance in ZS11, which will greatly promote the application of this superior trait in rapeseed breeding.

Genetic dissection of branch architecture in oilseed rape (Brassica napus L.) germplasm.

Branch architecture is an important factor influencing rapeseed planting density, mechanized harvest, and yield. However, its related genes and regulatory mechanisms remain largely unknown. In this study, branch angle (BA) and branch dispersion degree (BD) were used to evaluate branch architecture. Branch angle exhibited a dynamic change from an increase in the early stage to a gradual decrease until reaching a stable state. Cytological analysis showed that BA variation was mainly due to xylem size differences in the vascular bundle of the branch junction. The phenotypic analysis of 327 natural accessions revealed that BA in six environments ranged from 24.3° to 67.9°, and that BD in three environments varied from 4.20 cm to 21.4 cm, respectively. A total of 115 significant loci were detected through association mapping in three models (MLM, mrMLM, and FarmCPU), which explained 0.53%-19.4% of the phenotypic variations. Of them, 10 loci were repeatedly detected in different environments and models, one of which qBAD.A03-2 was verified as a stable QTL using a secondary segregation population. Totally, 1066 differentially expressed genes (DEGs) were identified between branch adaxial- and abaxial- sides from four extremely large or small BA/BD accessions through RNA sequencing. These DEGs were significantly enriched in the pathways related to auxin biosynthesis and transport as well as cell extension such as indole alkaloid biosynthesis, other glycan degradation, and fatty acid elongation. Four known candidate genes BnaA02g16500D (PIN1), BnaA03g10430D (PIN2), BnaC03g06250D (LAZY1), and BnaC06g20640D (ARF17) were identified by both GWAS and RNA-seq, all of which were involved in regulating the asymmetric distribution of auxins. Our identified association loci and candidate genes provide a theoretical basis for further study of gene cloning and genetic improvement of branch architecture.

Quantitative trait loci mapping reveals important genomic regions controlling root architecture and shoot biomass under nitrogen, phosphorus, and potassium stress in rapeseed (Brassica napus L.).

Plants rely on root systems for nutrient uptake from soils. Marker-assisted selection helps breeders to select desirable root traits for effective nutrient uptake. Here, 12 root and biomass traits were investigated at the seedling stage under low nitrogen (LN), low phosphorus (LP), and low potassium (LK) conditions, respectively, in a recombinant inbred line (RIL) population, which was generated from Brassica napus L. Zhongshuang11 and 4D122 with significant differences in root traits and nutrient efficiency. Significant differences for all the investigated traits were observed among RILs, with high heritabilities (0.43-0.74) and high correlations between the different treatments. Quantitative trait loci (QTL) mapping identified 57, 27, and 36 loci, explaining 4.1-10.9, 4.6-10.8, and 4.9-17.4% phenotypic variances under LN, LP, and LK, respectively. Through QTL-meta analysis, these loci were integrated into 18 significant QTL clusters. Four major QTL clusters involved 25 QTLs that could be repeatedly detected and explained more than 10% phenotypic variances, including two NPK-common and two specific QTL clusters (K and NK-specific), indicating their critical role in cooperative nutrients uptake of N, P, and K. Moreover, 264 genes within the four major QTL clusters having high expressions in roots and SNP/InDel variations between two parents were identified as potential candidate genes. Thirty-eight of them have been reported to be associated with root growth and development and/or nutrient stress tolerance. These key loci and candidate genes lay the foundation for deeper dissection of the NPK starvation response mechanisms in B. napus.

Deciphering the Genetic Basis of Root and Biomass Traits in Rapeseed (Brassica napus L.) through the Integration of GWAS and RNA-Seq under Nitrogen Stress.

An excellent root system is responsible for crops with high nitrogen-use efficiency (NUE). The current study evaluated the natural variations in 13 root- and biomass-related traits under a low nitrogen (LN) treatment in a rapeseed association panel. The studied traits exhibited significant phenotypic differences with heritabilities ranging from 0.53 to 0.66, and most of the traits showed significant correlations with each other. The genome-wide association study (GWAS) found 51 significant and 30 suggestive trait-SNP associations that integrated into 14 valid quantitative trait loci (QTL) clusters and explained 5.7-21.2% phenotypic variance. In addition, RNA sequencing was performed at two time points to examine the differential expression of genes (DEGs) between high and low NUE lines. In total, 245, 540, and 399 DEGs were identified as LN stress-specific, high nitrogen (HN) condition-specific, and HNLN common DEGs, respectively. An integrated analysis of GWAS, weighted gene co-expression network, and DEGs revealed 16 genes involved in rapeseed root development under LN stress. Previous studies have reported that the homologs of seven out of sixteen potential genes control root growth and NUE. These findings revealed the genetic basis underlying nitrogen stress and provided worthwhile SNPs/genes information for the genetic improvement of NUE in rapeseed.

Genome-Wide Association Studies of Root-Related Traits in Brassica napus L. under Low-Potassium Conditions.

Roots are essential organs for a plant's ability to absorb water and obtain mineral nutrients, hence they are critical to its development. Plants use root architectural alterations to improve their chances of absorbing nutrients when their supply is low. Nine root traits of a Brassica napus association panel were explored in hydroponic-system studies under low potassium (K) stress to unravel the genetic basis of root growth in rapeseed. The quantitative trait loci (QTL) and candidate genes for root development were discovered using a multilocus genome-wide association study (ML-GWAS). For the nine traits, a total of 453 significant associated single-nucleotide polymorphism (SNP) loci were discovered, which were then integrated into 206 QTL clusters. There were 45 pleiotropic clusters, and qRTA04-4 and qRTC04-7 were linked to TRL, TSA, and TRV at the same time, contributing 5.25-11.48% of the phenotypic variance explained (PVE) to the root traits. Additionally, 1360 annotated genes were discovered by examining genomic regions within 100 kb upstream and downstream of lead SNPs within the 45 loci. Thirty-five genes were identified as possibly regulating root-system development. As per protein-protein interaction analyses, homologs of three genes (BnaC08g29120D, BnaA07g10150D, and BnaC04g45700D) have been shown to influence root growth in earlier investigations. The QTL clusters and candidate genes identified in this work will help us better understand the genetics of root growth traits and could be employed in marker-assisted breeding for rapeseed adaptable to various conditions with low K levels.

Discovery of Genomic Regions and Candidate Genes Controlling Root Development Using a Recombinant Inbred Line Population in Rapeseed (Brassica napus L.).

Marker-assisted selection enables breeders to quickly select excellent root architectural variations, which play an essential role in plant productivity. Here, ten root-related and shoot biomass traits of a new F6 recombinant inbred line (RIL) population were investigated under hydroponics and resulted in high heritabilities from 0.61 to 0.83. A high-density linkage map of the RIL population was constructed using a Brassica napus 50k Illumina single nucleotide polymorphism (SNP) array. A total of 86 quantitative trait loci (QTLs) explaining 4.16-14.1% of the phenotypic variances were detected and integrated into eight stable QTL clusters, which were repeatedly detected in different experiments. The codominant markers were developed to be tightly linked with three major QTL clusters, qcA09-2, qcC08-2, and qcC08-3, which controlled both root-related and shoot biomass traits and had phenotypic contributions greater than 10%. Among these, qcA09-2, renamed RT.A09, was further fine-mapped to a 129-kb interval with 19 annotated genes in the B. napus reference genome. By integrating the results of real-time PCR and comparative sequencing, five genes with expression differences and/or amino acid differences were identified as important candidate genes for RT.A09. Our findings laid the foundation for revealing the molecular mechanism of root development and developed valuable markers for root genetic improvement in rapeseed.

Multi-Functional Development and Utilization of Rapeseed: Comprehensive Analysis of the Nutritional Value of Rapeseed Sprouts.

Rapeseed is the third largest oil crop in the world and the largest oil crop in China. The multi-functional development and utilization of rapeseed is an effective measure for the high-quality development of rapeseed industry in China. In this study, several basic nutrients of eight rapeseed sprouts and five bean sprouts (3-5 varieties each) were determined, including sugar, crude protein, crude fiber, vitamin E, minerals, fatty acids, amino acids, and glucosinolates. Data analysis revealed that compared with bean sprouts, rapeseed sprouts were nutritionally balanced and were richer in active nutrients such as glucose, magnesium, selenium, vitamin E, and glucosinolate. Moreover, rapeseed sprouts exhibited reasonable amino acid composition and abundant unsaturated fatty acids (accounting for 90.32% of the total fatty acids). All these results indicated the potential of rapeseed sprout as a functional vegetable. Subsequently, three dominant nutrients including vitamin E, glucosinolate, and selenium were investigated in seeds and sprouts of 44 B. napus L. varieties. The results showed that germination raised the ratio of α-tocopherol/γ-tocopherol from 0.53 in seeds to 9.65 in sprouts, greatly increasing the content of α-tocopherol with the strongest antioxidant activity among the eight isomers of vitamin E. Furthermore, germination promoted the conversion and accumulation of glucosinolate components, especially, glucoraphanin with strong anti-cancer activity with its proportion increased from 1.06% in seeds to 1.62% in sprouts. In addition, the contents of selenium, vitamin E, and glucosinolate in rapeseed sprouts were highly correlated with those in seeds. Furthermore, these three dominant nutrients varied greatly within B. napus varieties, indicating the great potential of rapeseed sprouts to be further bio-enhanced. Our findings provide reference for the multi-purpose development and utilization of rapeseed, lay a theoretical foundation for the development of rapeseed sprout into a functional vegetable, and provide a novel breeding direction.

Genetic Dissection of Mature Root Characteristics by Genome-Wide Association Studies in Rapeseed (Brassica napus L.).

Roots are complicated quantitative characteristics that play an essential role in absorbing water and nutrients. To uncover the genetic variations for root-related traits in rapeseed, twelve mature root traits of a Brassica napus association panel were investigated in the field within three environments. All traits showed significant phenotypic variation among genotypes, with heritabilities ranging from 55.18% to 79.68%. Genome-wide association studies (GWAS) using 20,131 SNPs discovered 172 marker-trait associations, including 103 significant SNPs (-log10 (p) > 4.30) that explained 5.24-20.31% of the phenotypic variance. With the linkage disequilibrium r2 > 0.2, these significant associations were binned into 40 quantitative trait loci (QTL) clusters. Among them, 14 important QTL clusters were discovered in two environments and/or with phenotypic contributions greater than 10%. By analyzing the genomic regions within 100 kb upstream and downstream of the peak SNPs within the 14 loci, 334 annotated genes were found. Among these, 32 genes were potentially associated with root development according to their expression analysis. Furthermore, the protein interaction network using the 334 annotated genes gave nine genes involved in a substantial number of interactions, including a key gene associated with root development, BnaC09g36350D. This research provides the groundwork for deciphering B. napus' genetic variations and improving its root system architecture.

Genome-wide association study and transcriptome analysis reveal key genes affecting root growth dynamics in rapeseed.

BACKGROUND: In terms of global demand, rapeseed is the third-largest oilseed crop after soybeans and palm, which produces vegetable oil for human consumption and biofuel for industrial production. Roots are vital organs for plant to absorb water and attain mineral nutrients, thus they are of great importance to plant productivity. However, the genetic mechanisms regulating root development in rapeseed remain unclear. In the present study, seven root-related traits and shoot biomass traits in 280 Brassica napus accessions at five continuous vegetative stages were measured to establish the genetic basis of root growth in rapeseed. RESULTS: The persistent and stage-specific genetic mechanisms were revealed by root dynamic analysis. Sixteen persistent and 32 stage-specific quantitative trait loci (QTL) clusters were identified through genome-wide association study (GWAS). Root samples with contrasting (slow and fast) growth rates throughout the investigated stages and those with obvious stage-specific changes in growth rates were subjected to transcriptome analysis. A total of 367 differentially expressed genes (DEGs) with persistent differential expressions throughout root development were identified, and these DEGs were significantly enriched in GO terms, such as energy metabolism and response to biotic or abiotic stress. Totally, 485 stage-specific DEGs with different expressions at specific stage were identified, and these DEGs were enriched in GO terms, such as nitrogen metabolism. Four candidate genes were identified as key persistent genetic factors and eight as stage-specific ones by integrating GWAS, weighted gene co-expression network analysis (WGCNA), and differential expression analysis. These candidate genes were speculated to regulate root system development, and they were less than 100 kb away from peak SNPs of QTL clusters. The homologs of three genes (BnaA03g52990D, BnaA06g37280D, and BnaA09g07580D) out of 12 candidate genes have been reported to regulate root development in previous studies. CONCLUSIONS: Sixteen QTL clusters and four candidate genes controlling persistently root development, and 32 QTL clusters and eight candidate genes stage-specifically regulating root growth in rapeseed were detected in this study. Our results provide new insights into the temporal genetic mechanisms of root growth by identifying key candidate QTL/genes in rapeseed.

Melatonin Represses Oil and Anthocyanin Accumulation in Seeds.

Previous studies have clearly demonstrated that the putative phytohormone melatonin functions directly in many aspects of plant growth and development. In Arabidopsis (Arabidopsis thaliana), the role of melatonin in seed oil and anthocyanin accumulation, and corresponding underlying mechanisms, remain unclear. Here, we found that serotonin N-acetyltransferase1 (SNAT1) and caffeic acid O-methyltransferase (COMT) genes were ubiquitously and highly expressed and essential for melatonin biosynthesis in Arabidopsis developing seeds. We demonstrated that blocking endogenous melatonin biosynthesis by knocking out SNAT1 and/or COMT significantly increased oil and anthocyanin content of mature seeds. In contrast, enhancement of melatonin signaling by exogenous application of melatonin led to a significant decrease in levels of seed oil and anthocyanins. Further gene expression analysis through RNA sequencing and reverse-transcription quantitative PCR demonstrated that the expression of a series of important genes involved in fatty acid and anthocyanin accumulation was significantly altered in snat1-1 comt-1 developing seeds during seed maturation. We also discovered that SNAT1 and COMT significantly regulated the accumulation of both mucilage and proanthocyanidins in mature seeds. These results not only help us understand the function of melatonin and provide valuable insights into the complicated regulatory network controlling oil and anthocyanin accumulation in seeds, but also divulge promising gene targets for improvement of both oil and flavonoids in seeds of oil-producing crops and plants.

Temporal genetic patterns of root growth in Brassica napus L. revealed by a low-cost, high-efficiency hydroponic system.

KEY MESSAGE: Application of a low-cost and high-efficiency hydroponic system in a rapeseed population verified two types of genetic factors ("persistent" and "stage-specific") that control root development. The root system is a vital plant component for nutrient and water acquisition and is targeted to enhance plant productivity. Genetic dissection of the root system generally focuses on a single stage, but roots grow continuously during plant development. To reveal the temporal genetic patterns of root development, we measured nine root-related traits in a rapeseed recombinant inbred line population at six continuous stages during vegetative growth, using a modified hydroponic system with low-cost and high-efficiency features that could synchronize plant growth under controlled conditions. Phenotypic correlation and growth dynamic analysis suggested the existence of two types of genetic factors ("persistent" and "stage-specific") that control root development. Dynamic (unconditional and conditional) quantitative trait loci (QTL) mapping detected 28 stage-specific and 23 persistent QTLs related to root growth. Among them, 13 early stage-specific, 19 persistent and 8 later stage-specific QTLs were detected at 7 DAS (days after sowing), 16 DAS and 5 EL (expanding leaf stage), respectively, providing efficient and adaptable stages for QTL identification. The effective prediction of biomass accumulation using root morphological traits (up to 96.6% or 92.64% at a specific stage or the final stage, respectively) verified that root growth allocation with maximum root uptake area facilitated biomass accumulation. Furthermore, marker-assistant selection, which combined the "persistent" and "stage-specific" QTLs, proved their effectiveness for root improvement with an excellent uptake area. Our results highlight the potential of high-throughput and precise phenotyping to assess the dynamic genetics of root growth and provide new insights into ideotype root system-based biomass breeding.

Genetic dissection of root morphological traits as related to potassium use efficiency in rapeseed under two contrasting potassium levels by hydroponics.

To reveal the genetic basis of potassium use efficiency (KUE) in rapeseed, root morphology (RM), biomass and KUE-related traits were measured in a recombinant inbred line population with 175 F7 lines that were subjected to high-potassium (HK) and low-potassium (LK) treatments by hydroponics. A total of 109 significant QTLs were identified to be associated with the examined traits. Sixty-one of these QTLs were integrated into nine stable QTLs. The higher heritability for RM and biomass traits and lower heritability for KUE-related traits, as well as nine stable QTLs for RM traits and only two for KUE-related traits, suggested that regulating RM traits would be more effective than selecting KUE traits directly to improve KUE by marker-assisted selection. Furthermore, the integration of stable QTLs identified in the HK, LK, high-nitrogen (HN) and low-nitrogen (LN) conditions gave 10 QTL clusters. Seven of these clusters were classified into major QTLs that explained 7.4%-23.7% of the total phenotypic variation. Five of the major QTL clusters were detected under all of the treated conditions, and four clusters were specifically detected under the LK and LN conditions. These common and specific QTL clusters may be useful for the simultaneous improvement of multiple traits by marker-assisted selection.

Genetic Dissection of Root Morphological Traits Related to Nitrogen Use Efficiency in Brassica napus L. under Two Contrasting Nitrogen Conditions.

As the major determinant for nutrient uptake, root system architecture (RSA) has a massive impact on nitrogen use efficiency (NUE). However, little is known the molecular control of RSA as related to NUE in rapeseed. Here, a rapeseed recombinant inbred line population (BnaZNRIL) was used to investigate root morphology (RM, an important component for RSA) and NUE-related traits under high-nitrogen (HN) and low-nitrogen (LN) conditions by hydroponics. Data analysis suggested that RM-related traits, particularly root size had significantly phenotypic correlations with plant dry biomass and N uptake irrespective of N levels, but no or little correlation with N utilization efficiency (NUtE), providing the potential to identify QTLs with pleiotropy or specificity for RM- and NUE-related traits. A total of 129 QTLs (including 23 stable QTLs, which were repeatedly detected at least two environments or different N levels) were identified and 83 of them were integrated into 22 pleiotropic QTL clusters. Five RM-NUE, ten RM-specific and three NUE-specific QTL clusters with same directions of additive-effect implied two NUE-improving approaches (RM-based and N utilization-based directly) and provided valuable genomic regions for NUE improvement in rapeseed. Importantly, all of four major QTLs and most of stable QTLs (20 out of 23) detected here were related to RM traits under HN and/or LN levels, suggested that regulating RM to improve NUE would be more feasible than regulating N efficiency directly. These results provided the promising genomic regions for marker-assisted selection on RM-based NUE improvement in rapeseed.

Comparative Transcriptome Analysis of Primary Roots of Brassica napus Seedlings with Extremely Different Primary Root Lengths Using RNA Sequencing.

Primary root (PR) development is a crucial developmental process that is essential for plant survival. The elucidation of the PR transcriptome provides insight into the genetic mechanism controlling PR development in crops. In this study, we performed a comparative transcriptome analysis to investigate the genome-wide gene expression profiles of the seedling PRs of four Brassica napus genotypes that were divided into two groups, short group (D43 and D61), and long group (D69 and D72), according to their extremely different primary root lengths (PRLs). The results generated 55,341,366-64,631,336 clean reads aligned to 62,562 genes (61.9% of the current annotated genes) in the B. napus genome. We provide evidence that at least 44,986 genes are actively expressed in the B. napus PR. The majority of the genes that were expressed during seedling PR development were associated with metabolism, cellular processes, response to stimulus, biological regulation, and signaling. Using a pairwise comparison approach, 509 differentially expressed genes (DEGs; absolute value of log2 fold-change ≥1 and p ≤ 0.05) between the long and short groups were revealed, including phytohormone-related genes, protein kinases and phosphatases, oxygenase, cytochrome P450 proteins, etc. Combining GO functional category, KEGG, and MapMan pathway analyses indicated that the DEGs involved in cell wall metabolism, carbohydrate metabolism, lipid metabolism, secondary metabolism, protein modification and degradation, hormone pathways and signaling pathways were the main causes of the observed PRL differences. We also identified 16 differentially expressed transcription factors (TFs) involved in PR development. Taken together, these transcriptomic datasets may serve as a foundation for the identification of candidate genes and may provide valuable information for understanding the molecular and cellular events related to PR development.

Altered Transcription and Neofunctionalization of Duplicated Genes Rescue the Harmful Effects of a Chimeric Gene in Brassica napus.

Chimeric genes contribute to the evolution of diverse functions in plants and animals. However, new chimeric genes also increase the risk of developmental defects. Here, we show that the chimeric gene Brassica napus male sterile 4 (Bnams4b ) is responsible for genic male sterility in the widely used canola line 7365A (Bnams3 ms3ms4bms4b ). Bnams4b originated via exon shuffling ∼4.6 million years ago. It causes defects in the normal functions of plastids and induces aborted anther formation and/or albino leaves and buds. Evidence of the age of the mutation, its tissue expression pattern, and its sublocalization indicated that it coevolved with BnaC9.Tic40 (BnaMs3). In Arabidopsis thaliana, Bnams4b results in complete male sterility that can be rescued by BnaC9.Tic40, suggesting that BnaC9.Tic40 might restore fertility through effects on protein level. Another suppressor gene, Bnams4a , rescues sterility by reducing the level of transcription of Bnams4b Our results suggest that Brassica plants have coevolved altered transcription patterns and neofunctionalization of duplicated genes that can block developmental defects resulting from detrimental chimeric genes.