Genome-Wide Identification, Expression Analysis and Functional Study of DELLA Genes in Chinese Cabbage (Brassica rapa L. ssp. pekinensis).

BACKGROUND: DELLA protein is a crucial factor which played pivotal roles in regulating numerous intriguing biological processes in plant development and abiotic stress responses. However, little is known about the function and information of DELLA protein in Chinese cabbage. METHODS: Using 5 DELLA gene sequences in Arabidopsis Thaliana as probes, 5 DELLA genes in Chinese cabbage were identified by Blast search in Chinese cabbage database (Brassica database (BRAD)). The National Center for Biotechnology Information (NCBI), ExPaSy, SWISS-MODEL, DNAMAN, MEGA 11, PlantCARE were used to identify and analyze the DELLA gene family of Chinese cabbage. Gene expression was analyzed by quantitative real-time polymerase chain reaction (qRT-PCR). The function of BraA10gRGL3 was verified by overexpression and phenotypic analysis of BraA10gRGL3 and yeast hybrid. RESULTS: In this study, 5 BraDELLAs homologous to Arabidopsis thaliana were identified and cloned based on the Brassica database, namely, BraA02gRGL1, BraA05gRGL2, BraA10gRGL3, BraA06gRGA and BraA09gRGA. All BraDELLAs contain the DELLA, TVHYNP, and GRAS conserved domains. Cis-element analysis revealed that the promoter regions of these 5 DELLA genes all contain light-responsive elements, TCT motif, I-box, G-box, and box 4, which are associated with GA signaling. Transcriptome analysis results proved that the expression of BraA02gRGL1, BraA05gRGL2, and BraA10gRGL3 in Y2 at different growth stages were lower than them in Y7, which is consistent with the phenotype that Y7 exhibited stronger stress tolerance than Y2. It is worth emphasizing that even through the overexpression of BraA10gRGL3-Y7 in Arabidopsis resulted in smaller leaf size and lower fresh weight compared to the wild type (WT) Arabidopsis: Columbia, a stronger response to abiotic stresses was observed in BraA10gRGL3-Y7. It indicated that BraA10gRGL3-Y7 can improve the stress resistance of plants by inhibiting their growth. Moreover, the yeast two-hybrid experiment confirmed that BraA10gRGL3-Y7 can interacted with BraA05gGID1a-Y7, BraA04gGID1b1, BraA09gGID1b3-Y2, and BraA06gGID1c, whereas BraA10gRGL3-Y2 cannot interact with any BraGID1. CONCLUSIONS: Collectively, BraDELLAs play important role in plant development and response to abiotic stress. The differences in amino acid sequences between BraA10gRGL3-Y2 and BraA10gRGL3-Y7 may result in variations in their protein binding sites, thus affecting their interaction with the BraGID1 family proteins. This systematic analysis lays the foundation for further study of the functional characteristics of DELLA genes of Chinese cabbage.

PHB3 interacts with BRI1 and BAK1 to mediate brassinosteroid signal transduction in Arabidopsis and tomato.

Brassinosteroids (BRs) are plant hormones that are essential in plant growth and development. BRASSINOSTEROID-INSENSITIVE 1 (BRI1) and BRI1 ASSOCIATED RECEPTOR KINASE 1 (BAK1), which are located on the plasma membrane, function as co-receptors that accept and transmit BR signals. PROHIBITIN 3 (PHB3) was identified in both BRI1 and BAK1 complexes by affinity purification and LC-MS/MS analysis. Biochemical data showed that BRI1/BAK1 interacted with PHB3 in vitro and in vivo. BRI1/BAK1 phosphorylated PHB3 in vitro. When the Thr-80 amino acid in PHB3 was mutated to Ala, the mutant protein was not phosphorylated by BRI1 and the mutant protein interaction with BRI1 was abolished in the yeast two-hybrid assay. BAK1 did not phosphorylate the mutant protein PHB3T54A . The loss-of-function phb3 mutant showed a weaker BR signal than the wild-type. Genetic analyses revealed that PHB3 is a BRI1/BAK1 downstream substrate that participates in BR signalling. PHB3 has five homozygous in tomato, and we named the closest to AtPHB3 as SlPHB3.1. Biochemical data showed that SlBRI1/SlSERK3A/SlSERK3B interacted with SlPHB3.1 and SlPHB3.3. The CRISPR-Cas9 method generated slphb3.1 mutant led to a BR signal stunted relatively in tomatoes. PHB3 is a new component of the BR signal pathway in both Arabidopsis and tomato.

Genome-Wide Identification of the NRT1 Family Members and Their Expression under Low-Nitrate Conditions in Chinese Cabbage (Brassica rapa L. ssp. pekinensis).

Nitrate transporters (NRTs) actively take up and transform nitrate (N) to form a large family with many members and distinct functions in plant growth and development. However, few studies have identified them in the context of low nitrate concentrations in Chinese cabbage (Brassica rapa L. ssp. Pekinensis), an important vegetable in China. This study focuses on the identification and analysis of the nitrate transporter 1 (NRT1) gene family as well as various aspects, including its phylogenic distribution, chromosomal position, gene structure, conserved motifs, and duplication pattern. Using bioinformatics methods, we identified and analyzed 84 BrNRT1 genes distributed on ten chromosomes. Furthermore, we conducted an analysis of the expression profile of the NRT1 gene in various tissues of Chinese cabbage exposed to varying nitrate concentrations. A phylogenetic analysis revealed that BrNRT1s members are distributed in six distinct groups. Based on an analysis of gene structure and conserved motifs, it can be inferred that BrNRT1 exhibits a generally conserved structural pattern. The promoters of BrNRT1 were discovered to contain moosefs (MFS) elements, suggesting their potential role in the regulation of NO3- transport across the cell membrane in Chinese cabbage. A transcriptome study and a subsequent RT-qPCR analysis revealed that the expression patterns of some BrNRT1 genes were distinct to specific tissues. This observation implies these genes may contribute to nitrate uptake and transport in various tissues or organs. The results offer fundamental insights into investigating the NRT1 gene family in Chinese cabbage. These results provide basic information for future research on the functional characterization of NRT1 genes in Chinese cabbage and the elucidation of the molecular mechanisms underlying low nitrogen tolerance in Chinese cabbage.

Integrative analysis of physiology, biochemistry and transcriptome reveals the mechanism of leaf size formation in Chinese cabbage (Brassica rapa L. ssp. pekinensis).

Introduction: The leaf, the main product organ, is an essential factor in determining the Chinese cabbage growth, yield and quality. Methods: To explore the regulatory mechanism of leaf size development of Chinese cabbage, we investigated the leaf size difference between two high-generation inbred lines of Chinese cabbage, Y2 (large leaf) and Y7 (small leaf). Furtherly, the transcriptome and cis-acting elements analyses were conducted. Results and Discussion: According to our results, Y2 exhibited a higher growth rate than Y7 during the whole growth stage. In addition, the significant higher leaf number was observed in Y2 than in Y7. There was no significant difference in the number of epidermal cells and guard cells per square millimeter between Y2 and Y7 leaves. It indicated that cell numbers caused the difference in leaf size. The measurement of phytohormone content confirmed that GA1 and GA3 mainly play essential roles in the early stage of leaf growth, and IPA and ABA were in the whole leaf growth period in regulating the cell proliferation difference between Y2 and Y7. Transcriptome analysis revealed that cyclins BraA09g010980.3C (CYCB) and BraA10g027420.3C (CYCD) were mainly responsible for the leaf size difference between Y2 and Y7 Chinese cabbage. Further, we revealed that the transcription factors BraA09gMYB47 and BraA06gMYB88 played critical roles in the difference of leaf size between Y2 and Y7 through the regulation of cell proliferation. Conclusion: This observation not only offers essential insights into understanding the regulation mechanism of leaf development, also provides a promising breeding strategy to improve Chinese cabbage yield.

Metabolite Profiling and Comparative Metabolomics Analysis of Jiaozhou Chinese Cabbage (Brassica rapa L. ssp. pekinensis) Planted in Different Areas.

BACKGROUND: Chinese cabbage (Brassica rapa L. ssp. pekinensis) is one of the most popular vegetables in China because of its taste and health benefits. The area of production has obvious effects on the quality of Chinese cabbage. However, metabolite profiling and variations in different production areas are still unclear. METHODS: Here, widely targeted metabolite analyses based on the ultra-high-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) approach were performed to study the metabolite profiling of Chinese cabbage planted in the Jiaozhou and Jinan areas. RESULTS: A total of 531 metabolites were detected, of which 529 were present in the Chinese cabbage from both areas, 108 were found to be chemicals related to Chinese traditional medicine, and 79 were found to correspond to at least one disease. Chinese cabbage is rich in nutritious substances such as lipids, phenolic acids, amino acids and derivatives, nucleotides and derivatives, organic acids, flavonoids, glucosinolates, saccharides, alcohols, and vitamins. Comparative analysis showed that the metabolic profiles differed between areas, and 89 differentially altered metabolites (DAMs) were characterized. Of these, 78 DAMs showed higher levels in Jinan Chinese cabbage, whereas 11 had higher levels in Jiaozhou Chinese cabbage. Two metabolites, S-(Methyl)glutathione and nicotinic acid adenine dinucleotide, were unique in Jiaozhou Chinese cabbage. Based on Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis, the DAMs were enriched into 23 pathways, of which tryptophan metabolism and thiamine metabolism were the significant enrichment pathways. CONCLUSIONS: This study provides new insights into the metabolite profiles and production areas affecting the metabolite variations of Chinese cabbage, which will be useful for functional Chinese cabbage cultivation.

Comparative Transcriptome and Co-Expression Network Analyses Reveal the Molecular Mechanism of Calcium-Deficiency-Triggered Tipburn in Chinese Cabbage (Brassica rapa L. ssp. Pekinensis).

Chinese cabbage tipburn is characterized by the formation of necrotic lesions on the margin of leaves, including on the insides of the leafy head. This physiological disorder is associated with a localized calcium deficiency during leaf development. However, little information is available regarding the molecular mechanisms governing Ca-deficiency-triggered tipburn. This study comprehensively analysed the transcriptomic comparison between control and calcium treatments (CK and 0 mM Ca) in Chinese cabbage to determine its molecular mechanism in tipburn. Our analysis identified that the most enriched gene ontology (GO) categories are photosynthesis, thylakoid and cofactor binding. Moreover, the KEGG pathway was most enriched in photosynthesis, carbon metabolism and carbon fixation. We also analyzed the co-expression network by functional categories and identified ten critical hub differentially expressed genes (DEGs) in each gene regulatory network (GRN). These DEGs might involve abiotic stresses, developmental processes, cell wall metabolism, calcium distribution, transcription factors, plant hormone biosynthesis and signal transduction pathways. Under calcium deficiency, CNX1, calmodulin-binding proteins and CMLs family proteins were downregulated compared to CK. In addition, plant hormones such as GA, JA, BR, Auxin and ABA biosynthesis pathways genes were downregulated under calcium treatment. Likewise, HATs, ARLs and TCP transcription factors were reported as inactive under calcium deficiency, and potentially involved in the developmental process. This work explores the specific DEGs' significantly different expression levels in 0 mM Ca and the control involved in plant hormones, cell wall developments, a light response such as chlorophylls and photosynthesis, transport metabolism and defence mechanism and redox. Our results provide critical evidence of the potential roles of the calcium signal transduction pathway and candidate genes governing Ca-deficiency-triggered tipburn in Chinese cabbage.

Identification of key genes controlling soluble sugar and glucosinolate biosynthesis in Chinese cabbage by integrating metabolome and genome-wide transcriptome analysis.

Introduction: Soluble sugar and glucosinolate are essential components that determine the flavor of Chinese cabbage and consumer preferences. However, the underlying regulatory networks that modulate the biosynthesis of soluble sugar and glucosinolate in Chinese cabbage remain largely unknown. Methods: The glucosinolate and carotene content in yellow inner-leaf Chinese cabbage were observed, followed by the combination of metabolome and transcriptome analysis to explore the metabolic basis of glucosinolate and soluble sugar. Results: This study observed high glucosinolate and carotene content in yellow inner-leaf Chinese cabbage, which showed a lower soluble sugar content. The differences between the yellow and the white inner-leaf Chinese cabbage were compared using the untargeted metabonomic and transcriptomic analyses in six cultivars of Chinese cabbage to explore the metabolic basis of glucosinolate and soluble sugar. Aliphatic glucosinolate and two soluble sugars (fructose and glucose) were the key metabolites that caused the difference in Chinese cabbage's glucosinolate and soluble sugar. By integrating soluble sugar and glucosinolate-associated metabolism and transcriptome data, we indicated BraA05gAOP1 and BraA04gAOP4, BraA03gHT7 and BraA01gHT4 were the glucosinolates and soluble sugar biosynthesis structural genes. Moreover, BraA01gCHR11 and BraA07gSCL1 were two vital transcription factors that regulate soluble sugar and glucosinolate biosynthesis. Discussion: These findings provide novel insights into glucosinolate and soluble sugar biosynthesis and a possible explanation for the significant difference in nutrients between yellow and white inner-leaf Chinese cabbage. Moreover, it will facilitate genetic modification to improve the Chinese cabbage's nutritional and health values.

Genetic and Transcriptome Analysis of Leaf Trichome Development in Chinese Cabbage (Brassica rapa L. subsp. pekinensis) and Molecular Marker Development.

Chinese cabbage (Brassica rapa L. subsp. pekinensis) is one of the vegetables with the largest cultivated area in China and has been a great addition to the daily diet of Chinese people. A genetic map has been constructed in our previous study using the F2 population of two inbred lines of Chinese cabbage, namely "G291" (a hairy line) and "ZHB" (a hairless line), based on which a candidate gene related to trichome traits was identified on chromosome A06 with a phenotypic variance of 47%. A molecular marker was found to co-segregate with the trichome traits of the F2 population, which is in the 5'-flanking region of BrGL1, and a corresponding patent has been granted (NO. CN 108545775 B). Transcriptome analysis was carried out on the cotyledon, the first true leaf and the leaf closest to each inflorescence of F2 individuals of "G291 × ZHB" with or without trichomes, respectively. Ten pathways, including 189 DEGs, were identified to be involved in the development of trichomes in Chinese cabbage, which may be specifically related to the development of leaf trichomes. Most of the pathways were related to the biosynthesis of the secondary metabolites, which may help plants to adapt to the ever-changing external environment. DEGs also enriched the "plant-pathogen interaction" pathway, which is consistent with the conclusion that trichomes are related to the disease resistance of plants. Our study provides a basis for future research on the occurrence and development of trichomes in Chinese cabbage.

Genome-Wide Identification and Characterization of Chinese Cabbage S1fa Transcription Factors and Their Roles in Response to Salt Stress.

The S1fa transcription factor is part of a small family involved in plant growth and development and abiotic stress tolerance. However, the roles of the S1fa genes in abiotic stress tolerance in Chinese cabbage are still unclear. In this study, four S1fa genes in the Chinese cabbage genome were identified and characterized for abiotic stress tolerance. Tissue-specific expression analysis suggested that three of these four S1fa genes were expressed in all tissues of Chinese cabbage, while Bra006994 was only expressed in the silique. Under Hg and Cd stresses, the S1fa genes were significantly expressed but were downregulated under NaCl stresses. The Bra034084 and Bra029784 overexpressing yeast cells exhibited high sensitivity to NaCl stresses, which led to slower growth compared with the wild type yeast cells (EV) under 1 M NaCl stress. In addition, the growth curve of the Bra034084 and Bra029784 overexpressing cells shows that the optical density was reduced significantly under salt stresses. The activities of the antioxidant enzymes, SOD, POD and CAT, were decreased, and the MDA, H2O2 and O2- contents were increased under salt stresses. The expression levels of cell wall biosynthesis genes Ccw14p, Cha1p, Cwp2p, Sed1p, Rlm1p, Rom2p, Mkk1p, Hsp12p, Mkk2p, Sdp1p and YLR194c were significantly enhanced, while Bck1p, and Ptc1p were downregulated under salt stresses. These results suggest that the Bra034084 and Bra029784 genes regulate cell wall biosynthesis and the defense regulatory system under salt stresses. These findings provide a fundamental basis for the further investigation of crop genetic modification to improve crop production and abiotic stress tolerance in Chinese cabbage.

Contrasting processing tomato cultivars unlink yield and pollen viability under heat stress.

Climate change is causing temperature increment in crop production areas worldwide, generating conditions of heat stress that negatively affect crop productivity. Tomato (Solanum lycopersicum), a major vegetable crop, is highly susceptible to conditions of heat stress. When tomato plants are exposed to ambient day/night temperatures that exceed 32 °C/20 °C, respectively, during the reproductive phase, fruit set and fruit weight are reduced, leading to a significant decrease in yield. Processing tomato cultivars are cultivated in open fields, where environmental conditions are not controlled; therefore, plants are exposed to multiple abiotic stresses, including heat stress. Nonetheless, information on stress response in processing tomatoes is very limited. Understanding the physiological response of modern processing tomato cultivars to heat stress may facilitate the development of thermotolerant cultivars. Here, we compared two tomato processing cultivars, H4107 and H9780, that we found to be constantly differing in yield performance. Using field and temperature-controlled greenhouse experiments, we show that the observed difference in yield is attributed to the occurrence of heat stress conditions. In addition, fruit set and seed production were significantly higher in the thermotolerant cultivar H4107, compared with H9780. Despite the general acceptance of pollen viability as a measure of thermotolerance, there was no difference in the percentage of viable pollen between H4107 and H9780 under either of the conditions tested. In addition to observations of similar pollen germination and bud abscission rates, our results suggest that processing tomato cultivars may present a particular case, in which pollen performance is not determining reproductive thermotolerance. Our results also demonstrate the value of combining controlled and uncontrolled experimental settings, in order to validate and identify heat stress-related responses, thus facilitating the development of thermotolerant processing tomato cultivars.

Enhanced Reproductive Thermotolerance of the Tomato high pigment 2 Mutant Is Associated With Increased Accumulation of Flavonols in Pollen.

Climate change has created an environment where heat stress conditions are becoming more frequent as temperatures continue to raise in crop production areas around the world. This situation leads to decreased crop production due to plant sensitivity to heat stress. Reproductive success is critically dependent on plants' ability to produce functional pollen grains, which are the most thermo-sensitive tissue. Flavonols are plant secondary metabolites known for their potent antioxidative activity, essential for male fertility in several species including tomato, and implicated in heat stress tolerance. Since flavonols are highly abundant in fruits of the tomato high pigment 2 (hp2) mutant, we tested the level of flavonols in pollen of this mutant, under the hypothesis that increased accumulation of flavonols would render pollen more tolerant to heat stress. Indeed, pollen from two alleles of the hp2 mutant was found to have flavonols levels increased by 18 and 280% compared with wild-type (WT) under moderate chronic heat stress (MCHS) conditions. This mutant produced on average 7.8-fold higher levels of viable pollen and displayed better germination competence under heat stress conditions. The percentage of fully seeded fruits and the number of seeds per fruit were maintained in the mutant under heat stress conditions while decreased in wild-type plants. Our results strongly suggest that increased concentrations of pollen flavonols enhance pollen thermotolerance and reproductive success under heat stress conditions. Thus, the high flavonols trait may help frame the model for improving crop resilience to heat stress.

Comparative transcriptome analysis between a resistant and a susceptible Chinese cabbage in response to Hyaloperonospora brassicae.

Downy mildew caused by Hyaloperonosporabrassicae (H. brassicae) leads to up to 90% of the crop yield loss in Chinese cabbage in China. A transcriptome analysis was carried out between a resistant line (13-13, R) and a susceptible line (15-14, S) of Chinese cabbage in response to H. brassicae. The NOISeq method was used to find differentially expressed genes (DEGs) between these two groups and GO and KEGG were carried out to find R genes related to downy mildew response of Chinese cabbage. qRT-PCR was carried out to verify the reliability of RNA-seq expression data. A total of 3,055 DEGs were screened out from 41,020 genes and clustered into 6 groups with distinct expression patterns. A total of 87 candidate DEGs were identified by functional annotation based on GO and KEGG analysis. These candidate genes are involved in plant-pathogen interaction pathway, among which 54 and 33 DEGs were categorized into plant-pathogen interaction proteins and transcription factors, respectively. Proteins encoded by these genes have been reported to play an important role in the pattern-triggered immunity (PTI) and effector-triggered immunity (ETI) processes of disease responses in some model plants, such as Arabidopsis, rice, tobacco, and tomato. However, little is known about the mechanisms of these genes in resistance to downy mildew in Chinese cabbage. Our findings are useful for further characterization of these candidate genes and helpful in breeding resistant strains.

Genome-Wide Identification and Analysis of the Cytochrome B5 Protein Family in Chinese Cabbage (Brassica rapa L. ssp. Pekinensis).

Cytochrome B5 (CB5) family proteins play an important role in various oxidation/reduction reactions in cells as the electron donor and are involved in a variety of biotic and abiotic stress processes. However, the function of the CB5s in Brassica rapa is still unclear. In this study, we carried out genome-wide identification, characterization, and expression analysis of BrCB5s in different tissues under adversities and stresses. It was identified that fifteen BrCB5s were distributed on different chromosomes, which were classified into seven groups (A-G) according to its phylogenetic relationship. Phylogenetic analysis of the CB5 protein sequences from six species showed that the BrCB5s conduct a close evolutionary process with the CB5s of Arabidopsis thaliana and far from those of Oryza sativa. Protein interaction analysis showed that 40 interaction patterns were predicted including two Sucrose Transporter 4 subfamily proteins (SUT 4) and Fatty Acid Hydroxylase 2 protein (FAH 2) can interact with most members of BrCB5s. The expression profile analysis indicated that BrCB5s were differentially expressed in different tissues, and the transcript abundances were significantly different under various abiotic stresses and plant hormone treatments. Our study provides a basis for a better understanding of the characteristics and biological functions of the CB5 family genes in Chinese cabbage during plant development, especially in plant responses to multiple stresses.

Ectopic expression of a Brassica rapa AINTEGUMENTA gene (BrANT-1) increases organ size and stomatal density in Arabidopsis.

The AINTEGUMENTA-like (AIL) family plays a central role in regulating the growth and development of organs in many plants. However, little is known about the characteristics and functions of the AIL family in Chinese cabbage (Brassica rapa L. ssp. pekinensis). In this study, a genome-wide analysis was performed to identify the members of the AIL family in Chinese cabbage. We identified three ANT genes and six ANT-like genes of Chinese cabbage, most of which were differentially expressed in different organs or tissues. Furthermore, compared with the wild-type line, the size of different organs in the 35S-BrANT-1 line was significantly increased by promoting cell proliferation. Meanwhile, over-expression of BrANT-1 also increases the stomatal number and delays the leaf senescence. Transcriptome analyses revealed that a set of cell proliferation and stoma development genes were up-regulated, while the senescence-associated genes were down-regulated, suggesting these genes may be involved in BrANT-1 regulated processes for controlling organ size, stomatal density and leaf senescence. In summary, this study offers important insights into the characteristics and functions of the ANT genes in Chinese cabbage, and provides a promising strategy to improve yield or head size in Chinese cabbage breeding programs.

Identification of miRNAs and their targets in regulating tuberous root development in radish using small RNA and degradome analyses.

High-throughput small RNA sequencing and degradome analysis were used in this study to thoroughly investigate the role of miRNA-mediated regulatory network in tuberous root development of radish. Samples from the early seedling stage (RE) and the cortex splitting stage (RL) were used for the construction of six small RNA libraries and one degradome library. A total of 518 known and 976 novel miRNAs were identified, of which, 338 known and 18 novel miRNAs were expressed in all six libraries, respectively. A total of 52 known and 57 novel miRNAs were identified to be significantly differentially expressed between RE and RL, and 195 mRNAs were verified to be the targets of 194 miRNAs by degradome sequencing. According to the degradome analysis, 11 differentially expressed miRNAs had miRNA-mRNA targets, and 13 targets were identified for these 11 miRNAs. Of the 13 miRNA-mRNA targets, 4 genes (RSG11079.t1, RSG11844.t1, RSG16775.t1, and RSG42419.t1) were involved in hormone-mediated signaling pathway, 2 gens (RSG11079.t1 and RSG16775.t1) were related to post-embryonic root development, and 1 gene (RSG23799.t1) was involved in anatomical structure morphogenesis, according to the GO function analysis for biological process. Target Genes participated in these processes are important candidates for further studies. This study provides valuable information for a better understanding of the molecular mechanisms involved in radish tuberous root formation and development.

Physiological and Transcriptomic Responses of Chinese Cabbage (Brassica rapa L. ssp. Pekinensis) to Salt Stress.

Salt stress is one of the major abiotic stresses that severely impact plant growth and development. In this study, we investigated the physiological and transcriptomic responses of Chinese cabbage "Qingmaye" to salt stress, a main variety in North China. Our results showed that the growth and photosynthesis of Chinese cabbage were significantly inhibited by salt treatment. However, as a glycophyte, Chinese cabbage could cope with high salinity; it could complete an entire life cycle at 100 mM NaCl. The high salt tolerance of Chinese cabbage was achieved by accumulating osmoprotectants and by maintaining higher activity of antioxidant enzymes. Transcriptomic responses were analyzed using the digital gene expression profiling (DGE) technique after 12 h of treatment by 200 mM NaCl. A total of 1235 differentially expressed genes (DEGs) including 740 up- and 495 down-regulated genes were identified. Functional annotation analyses showed that the DEGs were related to signal transduction, osmolyte synthesis, transcription factors, and antioxidant proteins. Taken together, this study contributes to our understanding of the mechanism of salt tolerance in Chinese cabbage and provides valuable information for further improvement of salt tolerance in Chinese cabbage breeding programs.

Transcriptome Analysis of Orange Head Chinese Cabbage (Brassica rapa L. ssp. pekinensis) and Molecular Marker Development.

Due to the visual appearance and high carotenoid content, orange inner leaves are a desirable trait for the Chinese cabbage. To understand the molecular mechanism underlying the formation of orange inner leaves, the BrCRTISO (Bra031539) gene, as the Br-or candidate gene, was analyzed among the white and orange varieties, and 7 single nucleotide polymorphisms (SNPs) were identified. However, only one SNP (C952 to T952) altered the amino acid sequence, resulting in a mutation from Leu318 to Phe318 in the orange varieties. Additionally, we analyzed differentially expressed genes (DEGs) between the orange and white F2 individuals (14-401 × 14-490) and found four downregulated genes were involved in the carotenoid biosynthesis pathway, which may lead to the accumulation of prolycopene and other carotenoid pigments in the orange inner leaves. In addition, we developed a novel InDel marker in the first intron, which cosegregates with the phenotypes of orange color inner leaves. In conclusion, these findings enhance our understanding of the underlying mechanism of pigment accumulation in the inner leaves of the Chinese cabbage. Additionally, the SNP (C952 to T952) and the InDel marker will facilitate the marker-assisted selection during Chinese cabbage breeding.

Comparative Transcriptome Analysis Reveals Effects of Exogenous Hematin on Anthocyanin Biosynthesis during Strawberry Fruit Ripening.

Anthocyanin in strawberries has a positive effect on fruit coloration. In this study, the role of exogenous hematin on anthocyanin biosynthesis was investigated. Our result showed that the white stage of strawberries treated with exogenous hematin had higher anthocyanin content, compared to the control group. Among all treatments, 5 µM of hematin was the optimal condition to promote color development. In order to explore the molecular mechanism of fruit coloring regulated by hematin, transcriptomes in the hematin- and non-hematin-treated fruit were analyzed. A large number of differentially expressed genes (DEGs) were identified in regulating anthocyanin synthesis, including the DEGs involved in anthocyanin biosynthesis, hormone signaling transduction, phytochrome signaling, starch and sucrose degradation, and transcriptional pathways. These regulatory networks may play an important role in regulating the color process of strawberries treated with hematin. In summary, exogenous hematin could promote fruit coloring by increasing anthocyanin content in the white stage of strawberries. Furthermore, transcriptome analysis suggests that hematin-promoted fruit coloring occurs through multiple related metabolic pathways, which provides valuable information for regulating fruit color via anthocyanin biosynthesis in strawberries.

Genome-Wide Identification and Analysis of the VQ Motif-Containing Protein Family in Chinese Cabbage (Brassica rapa L. ssp. Pekinensis).

Previous studies have showed that the VQ motif-containing proteins in Arabidopsis thaliana and Oryza sativa play an important role in plant growth, development, and stress responses. However, little is known about the functions of the VQ genes in Brassica rapa (Chinese cabbage). In this study, we performed genome-wide identification, characterization, and expression analysis of the VQ genes in Chinese cabbage, especially under adverse environment. We identified 57 VQ genes and classified them into seven subgroups (I-VII), which were dispersedly distributed on chromosomes 1 to 10. The expansion of these genes mainly contributed to segmental and tandem duplication. Fifty-four VQ genes contained no introns and 50 VQ proteins were less than 300 amino acids in length. Quantitative real-time PCR showed that the VQ genes were differentially expressed in various tissues and during different abiotic stresses and plant hormone treatments. This study provides a comprehensive overview of Chinese cabbage VQ genes and will benefit the molecular breeding for resistance to stresses and disease, as well as further studies on the biological functions of the VQ proteins.

Characterization and Development of EST-SSRs by Deep Transcriptome Sequencing in Chinese Cabbage (Brassica rapa L. ssp. pekinensis).

Simple sequence repeats (SSRs) are among the most important markers for population analysis and have been widely used in plant genetic mapping and molecular breeding. Expressed sequence tag-SSR (EST-SSR) markers, located in the coding regions, are potentially more efficient for QTL mapping, gene targeting, and marker-assisted breeding. In this study, we investigated 51,694 nonredundant unigenes, assembled from clean reads from deep transcriptome sequencing with a Solexa/Illumina platform, for identification and development of EST-SSRs in Chinese cabbage. In total, 10,420 EST-SSRs with over 12 bp were identified and characterized, among which 2744 EST-SSRs are new and 2317 are known ones showing polymorphism with previously reported SSRs. A total of 7877 PCR primer pairs for 1561 EST-SSR loci were designed, and primer pairs for twenty-four EST-SSRs were selected for primer evaluation. In nineteen EST-SSR loci (79.2%), amplicons were successfully generated with high quality. Seventeen (89.5%) showed polymorphism in twenty-four cultivars of Chinese cabbage. The polymorphic alleles of each polymorphic locus were sequenced, and the results showed that most polymorphisms were due to variations of SSR repeat motifs. The EST-SSRs identified and characterized in this study have important implications for developing new tools for genetics and molecular breeding in Chinese cabbage.