Comparative transcriptomic analysis unveils candidate genes associated with sugarcane growth rate.

Sugarcane (Saccharum spp.) growth is regulated by intricate gene networks and hormone secretions, positively correlating with sugarcane yield. There is a rising interest in exploring how the candidate genes found in sugarcane respond to plant growth. In this study, we simulated a typical growth environment to obtain accurate phenotypic data and screened for potential genes associated with plant growth through transcriptomics. Compared to Saccharum GuiTang 42, the other variety Saccharum GuiTang 44 exhibited earlier germination, a higher emergence rate, thicker pseudostems, taller plants, and a more extensive root system. The middle buds formed the greatest number of roots, followed by the lower and upper buds. Indole-3-acetic acid (IAA) and jasmonic acid effectively promoted bud development, while abscisic acid and trans-zeatin exhibited negative correlations with sugarcane bud growth. Transcriptome data from the upper, middle, and lower buds revealed 24,158 differentially expressed genes in all three comparisons, with MAPK signaling emerging as a critical pathway. The photosynthesis-antenna protein pathway is vital for middle and lower bud development during root germination. Lastly, key gene modules related to differences in hormone content between the two varieties were defined through weighted correlation network analysis and identified. The module significantly associated with IAA was enriched in pathways such as Proteasome and Protein processing in the endoplasmic reticulum, and the upregulation of key genes involved in this gene module had a highly significant positive correlation with bud outgrowth combined with IAA secretion. In conclusion, we have elucidated the pathways of hormones during sugarcane growth and the interactions between IAA and critical genes. These in-depth findings may guide modern sugarcane breeding.

MicroSugar: A database of comprehensive miRNA target prediction framework for sugarcane (Saccharum officinarum L.).

microRNA (miRNA) is a group of small non-coding RNA that plays important role in post-transcription of gene expression. With the studies about miRNA increase in sugarcane, the researchers lack an exhaustive resource to achieve the data. To fill this gap, we developed MicroSugar, a database that supported mRNA and miRNA annotation for sugarcane (http://suc.gene-db.com). MicroSugar is an integrated resource developed for 194,528 genes including 80,746 unigenes from long reads of Pacbio platform and 468 miRNAs from 72 samples. Internode elongation (jointing) is the key biological characteristic for the growth of sugarcane tillers into sugarcane stems. The present study combined the sequencing data from the different stages in internode elongation of stem and tiller. In total, the 14,300 3' untranslated region (UTR) sequences were extracted from the gene sequences and 3019 mRNAs as target of 327 miRNA were identified by miRanda algorithm and Spearman's Rho of expression levels. To determine the gene functions regulated by these miRNAs, the gene ontology enrichment analysis was performed and it confirmed that the over-represented Gene Ontology (GO) terms were associated with organism formation indicating the growth controlling function by miRNAs in sugarcane. Moreover, MicroSugar is a comprehensive and integrated database with a user-friendly responsive template. By browsing, searching and downloading of the nucleotide sequences, expression and miRNA targets, the user can retrieve information promptly. The database provides a valuable resource to facilitate the understanding of miRNA in sugarcane development and growth which will contribute to the study of sugarcane and other plants.

Transcriptome and metabolome analyses reveal new insights into chlorophyll, photosynthesis, metal ion and phenylpropanoids related pathways during sugarcane ratoon chlorosis.

BACKGROUND: Ratoon sugarcane is susceptible to chlorosis, characterized by chlorophyll loss, poor growth, and a multitude of nutritional deficiency mainly occurring at young stage. Chlorosis would significantly reduce the cane production. The molecular mechanism underlying this phenomenon remains unknown. We analyzed the transcriptome and metabolome of chlorotic and non-chlorotic sugarcane leaves of the same age from the same field to gain molecular insights into this phenomenon. RESULTS: The agronomic traits, such as plant height and the number of leaf, stalk node, and tillers declined in chlorotic sugarcane. Chlorotic leaves had substantially lower chlorophyll content than green leaves. A total of 11,776 differentially expressed genes (DEGs) were discovered in transcriptome analysis. In the KEGG enriched chlorophyll metabolism pathway, sixteen DEGs were found, eleven of which were down-regulated. Two photosynthesis pathways were also enriched with 32 genes downregulated and four genes up-regulated. Among the 81 enriched GO biological processes, there were four categories related to metal ion homeostasis and three related to metal ion transport. Approximately 400 metabolites were identified in metabolome analysis. The thirteen differentially expressed metabolites (DEMs) were all found down-regulated. The phenylpropanoid biosynthesis pathway was enriched in DEGs and DEMs, indicating a potentially vital role for phenylpropanoids in chlorosis. CONCLUSIONS: Chlorophyll production, metal ion metabolism, photosynthesis, and some metabolites in the phenylpropanoid biosynthesis pathway were considerably altered in chlorotic ratoon sugarcane leaves. Our finding revealed the relation between chlorosis and these pathways, which will help expand our mechanistic understanding of ratoon sugarcane chlorosis.

Global transcriptome changes of elongating internode of sugarcane in response to mepiquat chloride.

BACKGROUND: Mepiquat chloride (DPC) is a chemical that is extensively used to control internode growth and create compact canopies in cultured plants. Previous studies have suggested that DPC could also inhibit gibberellin biosynthesis in sugarcane. Unfortunately, the molecular mechanism underlying the suppressive effects of DPC on plant growth is still largely unknown. RESULTS: In the present study, we first obtained high-quality long transcripts from the internodes of sugarcane using the PacBio Sequel System. A total of 72,671 isoforms, with N50 at 3073, were generated. These long isoforms were used as a reference for the subsequent RNA-seq. Afterwards, short reads generated from the Illumina HiSeq 4000 platform were used to compare the differentially expressed genes in both the DPC and the control groups. Transcriptome profiling showed that most significant gene changes occurred after six days post DPC treatment. These genes were related to plant hormone signal transduction and biosynthesis of several metabolites, indicating that DPC affected multiple pathways, in addition to suppressing gibberellin biosynthesis. The network of DPC on the key stage was illustrated by weighted gene co-expression network analysis (WGCNA). Among the 36 constructed modules, the top positive correlated module, at the stage of six days post spraying DPC, was sienna3. Notably, Stf0 sulfotransferase, cyclin-like F-box, and HOX12 were the hub genes in sienna3 that had high correlation with other genes in this module. Furthermore, the qPCR validated the high accuracy of the RNA-seq results. CONCLUSION: Taken together, we have demonstrated the key role of these genes in DPC-induced growth inhibition in sugarcane.

Characterization of full-length transcriptome in Saccharum officinarum and molecular insights into tiller development.

BACKGROUND: Although extensive breeding efforts are ongoing in sugarcane (Saccharum officinarum L.), the average yield is far below the theoretical potential. Tillering is an important component of sugarcane yield, however, the molecular mechanism underlying tiller development is still elusive. The limited genomic data in sugarcane, particularly due to its complex and large genome, has hindered in-depth molecular studies. RESULTS: Herein, we generated full-length (FL) transcriptome from developing leaf and tiller bud samples based on PacBio Iso-Seq. In addition, we performed RNA-seq from tiller bud samples at three developmental stages (T0, T1 and T2) to uncover key genes and biological pathways involved in sugarcane tiller development. In total, 30,360 and 20,088 high-quality non-redundant isoforms were identified in leaf and tiller bud samples, respectively, representing 41,109 unique isoforms in sugarcane. Likewise, we identified 1063 and 1037 alternative splicing events identified in leaf and tiller bud samples, respectively. We predicted the presence of coding sequence for 40,343 isoforms, 98% of which was successfully annotated. Comparison with previous FL transcriptomes in sugarcane revealed 2963 unreported isoforms. In addition, we characterized 14,946 SSRs from 11,700 transcripts and 310 lncRNAs. By integrating RNA-seq with the FL transcriptome, 468 and 57 differentially expressed genes (DEG) were identified in T1vsT0 and T2vsT0, respectively. Strong up-regulation of several pyruvate phosphate dikinase and phosphoenolpyruvate carboxylase genes suggests enhanced carbon fixation and protein synthesis to facilitate tiller growth. Similarly, up-regulation of linoleate 9S-lipoxygenase and lipoxygenase genes in the linoleic acid metabolism pathway suggests high synthesis of key oxylipins involved in tiller growth and development. CONCLUSIONS: Collectively, we have enriched the genomic data available in sugarcane and provided candidate genes for manipulating tiller formation and development, towards productivity enhancement in sugarcane.

Integrated mRNA and small RNA sequencing reveals microRNA regulatory network associated with internode elongation in sugarcane (Saccharum officinarum L.).

BACKGROUND: Internode elongation is one of the most important traits in sugarcane because of its relation to crop productivity. Understanding the microRNA (miRNA) and mRNA expression profiles related to sugarcane internode elongation would help develop molecular improvement strategies but they are not yet well-investigated. To identify genes and miRNAs involved in internode elongation, the cDNA and small RNA libraries from the pre-elongation stage (EI), early elongation stage (EII) and rapid elongation stage (EIII) were sequenced and their expression were studied. RESULTS: Based on the sequencing results, 499,495,518 reads and 80,745 unigenes were identified from stem internodes of sugarcane. The comparisons of EI vs. EII, EI vs. EIII, and EII vs. EIII identified 493, 5035 and 3041 differentially expressed genes, respectively. Further analysis revealed that the differentially expressed genes were enriched in the GO terms oxidoreductase activity and tetrapyrrole binding. KEGG pathway annotation showed significant enrichment in "zeatin biosynthesis", "nitrogen metabolism" and "plant hormone signal transduction", which might be participating in internode elongation. miRNA identification showed 241 known miRNAs and 245 novel candidate miRNAs. By pairwise comparison, 11, 42 and 26 differentially expressed miRNAs were identified from EI and EII, EI and EIII, and EII and EIII comparisons, respectively. The target prediction revealed that the genes involved in "zeatin biosynthesis", "nitrogen metabolism" and "plant hormone signal transduction" pathways are targets of the miRNAs. We found that the known miRNAs miR2592-y, miR1520-x, miR390-x, miR5658-x, miR6169-x and miR8154-x were likely regulators of genes with internode elongation in sugarcane. CONCLUSIONS: The results of this study provided a global view of mRNA and miRNA regulation during sugarcane internode elongation. A genetic network of miRNA-mRNA was identified with miRNA-mediated gene expression as a mechanism in sugarcane internode elongation. Such evidence will be valuable for further investigations of the molecular regulatory mechanisms underpinning sugarcane growth and development.

A Zinc Oxide Nanoflower-Based Electrochemical Sensor for Trace Detection of Sunset Yellow.

Zinc oxide nanoflower (ZnONF) was synthesized by a simple process and was used to construct a highly sensitive electrochemical sensor for the detection of sunset yellow (SY). Due to the large surface area and high accumulation efficiency of ZnONF, the ZnONF-modified carbon paste electrode (ZnONF/CPE) showed a strong enhancement effect on the electrochemical oxidation of SY. The electrochemical behaviors of SY were investigated using voltammetry with the ZnONF-based sensor. The optimized parameters included the amount of ZnONF, the accumulation time, and the pH value. Under optimal conditions, the oxidation peak current was linearly proportional to SY concentration in the range of 0.50-10 µg/L and 10-70 µg/L, while the detection limit was 0.10 µg/L (signal-to-noise ratio = 3). The proposed method was used to determine the amount of SY in soft drinks with recoveries of 97.5%-103%, and the results were in good agreement with the results obtained by high-performance liquid chromatography.

Enhanced Activity of Genes Associated With Photosynthesis, Phytohormone Metabolism and Cell Wall Synthesis Is Involved in Gibberellin-Mediated Sugarcane Internode Growth.

Internode elongation is an important trait in sugarcane as it affects the sugarcane yield. Gibberellin (GA) is a key modulator of internode elongation in sugarcane. Understanding the gene expression features of GA-mediated internode elongation has both scientific and practical significance. This study aimed to examine the transcriptomic changes in the internode elongation of sugarcane following GA treatment. Eighteen cDNA libraries from the internode tissues on days of 0, 3, and 6 in control and GA treatment groups were sequenced and their gene expression were studied. RNA-seq analysis revealed 1,338,723,248 reads and 70,821 unigenes from elongating internodes of sugarcane. Comparative studies discovered a large number of transcripts that were differentially expressed in GA-treated samples compared to the control. Further analysis revealed that the differentially expressed genes were enriched in the metabolic process, one-carbon compound transport, and single-organism process. Kyoto Encyclopedia of Genes and Genomes pathway annotation showed significant enrichment in photosynthesis and plant hormone signal transduction, indicating its involvement in internode elongation. The function analysis suggested that metabolic pathways and biosynthesis of secondary metabolites, plant hormones, and cell wall components were enriched in the internodes of the GA-treated plants. The hub genes were identified, with the function of cellulose synthesis. The results of this study provide a global view of mRNA changes during sugarcane internode elongation and extend our knowledge of the GA-mediated cellular processes involved in sugarcane stem growth.

Genome-Wide Analysis of the DREB Subfamily in Saccharum spontaneum Reveals Their Functional Divergence During Cold and Drought Stresses.

Drought and cold stresses are the main environmental factors that affect the yield of sugarcane, and DREB genes play very important roles in tolerance to drought, cold, and other environmental stresses. In this study, bioinformatics analysis was performed to characterize Saccharum spontaneum SsDREB genes. RNA sequencing (RNA-seq) was used to detect the expression profiles of SsDREBs induced by cold and drought stresses. According to our results, there are 110 SsDREB subfamily proteins in S. spontaneum, which can be classified into six groups; 106 of these genes are distributed among 29 chromosomes. Inter- and intraspecies synteny analyses suggested that all DREB groups have undergone gene duplication, highlighting the polyploid events that played an important role in the expansion of the DREB subfamily. Furthermore, RNA-seq results showed that 45 SsDREBs were up- or downregulated under cold stress; 35 of them were found to be involved in responding to drought stress. According to protein-protein interaction analysis, SsDREB100, SsDREB102, and SsDREB105 play key roles during the response to cold stress. These results reveal that functional divergence exists between collinear homologous genes or among common origin genes in the DREB subfamily of S. spontaneum. This study presents a comprehensive analysis and systematic understanding of the precise mechanism of SsDREBs in response to abiotic stress and will lead to improvements in sugarcane.

Highly sensitive determination of capsaicin using a carbon paste electrode modified with amino-functionalized mesoporous silica.

Mesoporous silica (MS) and amino-functionalized mesoporous silica (NH(2)-FMS) were prepared and characterized using the techniques of transmission electron microscopy (TEM) and nitrogen adsorption-desorption. Voltammetry was used to investigate the electrochemical behavior of capsaicin at the amino-functionalized mesoporous silica, which was modified through carbon paste electrode (NH(2)-FMS/CPE). NH(2)-FMS/CPE showed better performance for the electrochemical oxidation of capsaicin, when compared with bare carbon paste electrode (CPE) and mesoporous silica modified carbon paste electrode (MS/CPE). We optimized the experimental conditions influencing the determination of capsaicin. Under optimal conditions, the oxidation peak current was proportional to capsaicin concentration in the range of 0.040-0.40 µmol L(-1)and 0.40-4.0 µmol L(-1), when the detection limit was 0.020 µmol L(-1) (S/N=3). The above method was successfully applied to determine capsaicin in hot pepper samples, yielding satisfactory results. The spiked recoveries were in the range of 93.1-100.7%.