Gene co-expression modules behind the three-pistil formation in wheat.

Multi-pistil trait in wheat is of great potential value in plant development research and crop breeding. Our previous studies identified the Pis1 locus that causes three pistils in wheat by genetic mapping using multiple DNA marker systems. However, there are still 26 candidate genes on the locus, and the causal gene remains to be found. In this study, we aimed to approach the molecular mechanism of multi-pistil formation. Comparative RNA sequencing (RNA-Seq) during the pistil formation was undertaken in four wheat lines: a three-pistil mutant TP, a single-pistil TILLING mutant of TP (SP), a three-pistil near-isogenic line CM28TP with the background of cultivar Chunmai 28 (CM28), and CM28. Electron microscopic analysis specified probable developmental stages of young spikes for the three-pistil formation. mRNA sequencing in the young spikes of the four lines represented 253 down-regulated genes and 98 up-regulated genes in both three-pistil lines, which included six potential genes for ovary development. Weighted gene co-expression analysis represented three-pistil trait-associated transcription factor-like genes, among which one hub gene, ARF5, was the most highlighted. ARF5 is on the Pis1 locus and an orthologue of MONOPTEROS which mediates tissue development in Arabidopsis. qRT-PCR validation implies that the deficiency of ARF5 underlies the three-pistil formation in wheat.

The overexpression of SlBRI1 driven by Atrd29A promoter-transgenic plants improves the chilling stress tolerance of tomato.

MAIN CONCLUSION: Overexpression of SlBRI1 driven by the Atrd29A promoter could increase the cold resistance of tomato plants during chilling stress but did not improve the expression of SlBRI1 and plant growth under normal conditions. Low temperature is the main limiting factor severely affecting tomato plant development, growth, and fruit quality in winter and spring. Brassinosteroids (BRs) and key BR signaling genes positively regulate tomato plant development and response to chilling stress. Brassinosteroid-insensitive 1 (BRI1) is a major BR receptor that initiates BR signaling. Our results showed that overexpression of SlBRI1 driven by the Atrd29A promoter in transgenic plants did not increase the expression of SlBRI1 under normal conditions but rapidly induced the expression of SlBRI1 during chilling stress. The degree of wilting was lower in Atrd29A promoter-transgenic plants than in wild-type (WT) plants after chilling stress. Atrd29A promoter-transgenic plants exhibited low relative electrolyte leakage and reactive oxygen species (ROS) accumulation under chilling stress. Transgenic plants showed higher photosynthetic ability and antioxidant enzyme activity than WT plants under chilling stress. The BR content and expression levels of key genes involved in BR biosynthesis in Atrd29A-promoter transgenic plants were significantly lower than those in WT plants during chilling stress. The abscisic acid (ABA) content and expression levels of key ABA biosynthesis genes in the Atrd29A promoter-transgenic plants were significantly higher than those in the WT plants during chilling stress. In addition, Atrd29A promoter-transgenic plants positively enhanced the expression levels of ICE-CBF-COR cold-responsive pathway genes. Therefore, the overexpression of SlBRI1 driven by the Atrd29A promoter in transgenic plants can be a valuable tool for reducing chilling stress.

Inheritance and QTL Mapping for Flower Color in Salvia miltiorrhiza Bunge.

Salvia miltiorrhiza Bunge is an outcross-pollinated plant with diverse flower colors, ranging from white to purple. To clarify the genetic basis of S. miltiorrhiza flower color, we crossed white-flowered S. miltiorrhiza f. alba with dark violet-flowered S. miltiorrhiza, and selfed F1 to obtain an F2 population. The RGB color system was used to describe the flower color of the parents, F1 progeny, and F2 individuals. Afterward, we used genotyping-by-sequencing technology to construct a high-density linkage map of S. miltiorrhiza based on the F2 population. Finally, the linkage map was used to locate the QTLs of the genes that control flower color in S. miltiorrhiza. Through measurement and cluster analysis of the R, G, and B values of flowers from the parents, F1, and F2 individuals, it was found that the purple flower color of S. miltiorrhiza is a quantitative trait controlled by 2 loci of major genes. The genetic map contained 605 SNPs with a total length of 738.3 cM in 8 linkage groups (LGs), and the average distance between 2 markers was 1.22 cM. Based on the constructed genetic map and the flower R, G, B, and R+G+B values, 2 QTLs were detected for flower color, located on LG4 and LG5. The results of this study lay the foundation for cloning genes that control flower color and studying the molecular mechanism of flower color regulation in S. miltiorrhiza.

HaCYC2c regulating the heteromorphous development and functional differentiation of florets by recognizing HaNDUA2 in sunflower.

KEY MESSAGE: The overexpression of HaCYC2c and its regulation on HaNDUA2 through transcriptional recognition are important for regulating the heteromorphous development and functional differentiation of ray and disc florets in sunflower. Flower symmetry is closely related to pollinator recruitment and individual fecundity for higher plants and is the main feature used to identify flower type in angiosperms. In sunflower, HaCYC2c regulates floral organ development and floral symmetry, but the specific detail remains unclear. In this study, sunflower long petal mutant (lpm) with HaCYC2c insertion mutation was used to investigate the regulating role of HaCYC2c in the morphogenesis of florets and the transformation of floral symmetry through phenotype, transcriptome, qRT-PCR, and possible protein-gene interactions analyses. Results showed that HaCYC2c was overexpressed after an insertion into the promoter region. This gene could recognize the cis-acting element GGTCCC in the promoter region of HaNDUA2 that might regulate HaNDUA2 and affect other related genes. As a consequence, the abnormal elongation of disc petals and the degradation of male reproductive system occurred at the early development of floral organ in sunflower. Furthermore, this insertion mutation resulted in floral symmetry transformation, from actinomorphy to zygomorphy, thereby making the tubular disc florets transformed into ray-like disc florets in sunflower lpm. The findings suggested that the overexpression of HaCYC2c and its control of HaNDUA2 through transcriptional recognition might be an important regulating node of the heteromorphous development and functional differentiation for ray and disc florets in sunflower. This node contributes to the understanding of the balance between pollinator recruitment capacity of ray florets and fertility of disc florets for the optimization of reproductive efficiency and enhancement of species competitiveness in sunflower.

Different responses to joint exposure to cadmium and zinc depends on the sex in Populus cathayana.

The alarming increase in soil contamination by heavy metals, such as cadmium and zinc demands immediate attention. The dioecious tree Populus cathayana, a phytoremediation plant, plays an important role in rehabilitating heavy metal contaminated areas. In this study, male and female P. cathayana plants were treated with Cd (20 mg kg-1) and different levels of Zn (25, 50, or 100 mg kg-1) to study their physiological responses. The results showed that Cd exposure alone caused stress by inhibiting the growth of both male and female plants. In both males and females, photosynthesis and antioxidant enzymes activities decreased substantially under Cd stress alone. Cd was largely located in the roots, but Zn was present in the shoots of both sexes. Zn supplementation considerably increased the photosynthetic rate from 14.62 % to 60.45 % and also enhanced the antioxidant enzymes activities from 24.11 % to 86.21 %. Zn treatment decreased the translocation ability of Cd compared to the Cd-only treatment, alleviating Cd toxicity. In addition, when sufficient Zn was made available, males showed a high degree of Cd accumulation, low root-to-shoot translocation, elevated antioxidant defense abilities, and an increased photosynthetic rate, while females were less responsive to Cd stress than males. Thus, combined exposure to Cd and Zn caused differential responses in plant growth and physiological processes between males and females P. cathayana. Male plants exhibit better Cd tolerance and accumulation capacity under optimum Zn supplementation. This study increases the fundamental knowledge regarding P. cathayana plants, which can be applied to enhance their remediation capacity in Cd-contaminated soils.

Improved cell seeding efficiency and cell distribution in porous hydroxyapatite scaffolds by semi-dynamic method.

Tissue engineering is a promising technique for the repair of bone defects. An efficient and homogeneous distribution of cell seeding into scaffold is a crucial but challenging step in the technique. Murine bone marrow mesenchymal stem cells were seeded into porous hydroxyapatite scaffolds of two morphologies by three methods: static seeding, semi-dynamic seeding, or dynamic perfusion seeding. Seeding efficiency, survival, distribution, and proliferation were quantitatively evaluated. To investigate the performance of the three seeding methods for larger/thicker scaffolds as well as batch seeding of numerous scaffolds, three scaffolds were stacked to form assemblies, and seeding efficiencies and cell distribution were analyzed. The semi-dynamic seeding and static seeding methods produced significantly higher seeding efficiencies, vitalities, and proliferation than did the dynamic perfusion seeding. On the other hand, the semi-dynamic seeding and dynamic perfusion seeding methods resulted in more homogeneous cell distribution than did the static seeding. For stacked scaffold assemblies, the semi-dynamic seeding method also created superior seeding efficiency and longitudinal cell distribution homogeneity. The semi-dynamic seeding method combines the high seeding efficiency of static seeding and satisfactory distribution homogeneity of dynamic seeding while circumventing their disadvantages. It may contribute to improved outcomes of bone tissue engineering.

TaEPFL1, an EPIDERMAL PATTERNING FACTOR-LIKE (EPFL) secreted peptide gene, is required for stamen development in wheat.

Members of the EPIDERMAL PATTERNING FACTOR-LIKE (EPFL) family play diverse roles in plant growth and development, including the guidance of inflorescence architecture and pedicel length. In this work, we identified and characterized the EFPL gene TaEPFL1 from the wheat pistillody mutant HTS-1. Sequence alignment and phylogenetic analysis indicated that TaEPFL1 belongs to the EPFL1 gene. Quantitative real-time RT-PCR analysis showed that the TaEPFL1 gene is expressed at an abnormally high level in pistillody stamens compared with that in pistils and stamens. Heterologous expression of the TaEPFL1 gene in Arabidopsis caused shortened filaments and pedicels and might reduce the level of AtACO2 gene expression. These results suggest that TaEPFL1 plays an important role in the development of stamen and that overexpression of TaEPFL1 results in abnormal stamens. We deduced that the overexpression of the TaEPFL1 gene may contribute to the homeotic transformation of stamens into pistils or pistil-like structures in wheat. These data offer insights into the molecular mechanism of pistillody mutation in wheat.

A base-promoted cascade reaction of α,ß-unsaturated N-tosylhydrazones with o-hydroxybenzyl alcohols: highly regioselective synthesis of N-sec-alkylpyrazoles.

An efficient method for the synthesis of N-sec-alkylpyrazoles through a base-promoted cascade cyclization/Michael addition reaction of α,ß-unsaturated N-tosylhydrazones with ortho-hydroxybenzyl alcohols has been developed. The desired products containing di- or triaryl groups at the same carbon atom were afforded in good to excellent yields with excellent regioselectivities (>20 : 1). Moreover, a three-component reaction of ortho-hydroxybenzyl alcohols, α,ß-unsaturated N-tosylhydrazones and saturated N-tosylhydrazones also took place to afford pyrazoles in good yields. This reaction offers a new route to triarylmethanes with a simple operation and is applicable for large-scale synthesis.

High-density genetic map construction and mapping of the homologous transformation sterility gene (hts) in wheat using GBS markers.

BACKGROUND: Homologous transformation sterility-1 (HTS-1) is a novel wheat mutant that exhibits pistillody, the transformation of stamens into pistils or pistil-like structures. More extreme phenotypes of this mutation can have six pistils or pistil-like structures without any stamens in a floret. Thus, HTS-1 is highly valuable for studies of wheat hybrid breeding and flower development. Previous studies have shown that two major genes (Pis1 and hts) control pistillody in HTS-1. The Pis1 gene controls the three-pistil trait in the three-pistil wheat mutant and has been mapped on chromosome 2D, but the hts gene has not been mapped or identified. To do so, we crossed HTS-1 with CM28TP (three-pistil mutant) and constructed a high-density linkage map with the F2 population (200 individuals). RESULTS: The map covered 2779.96 cM, and the genetic distance per chromosome ranged from 37.59 cM to 318.95 cM. The average distance between markers was 1.04 cM. We then mapped hts between GBS-SNP markers 4A_109 and 4A_119, separated by 2.0 cM and 5.2 Mb. To find the candidate genes, the hts region was enlarged to 7.2 Mb, encompassing 752 protein-coding genes. We identified TaWin1 as a possible candidate gene after comparing the 752 genes with 206 common differentially expressed genes between pistillody stamens (PS) versus normal stamens (S) and pistils (P) versus S. Real-time PCR indicated that TaWin1 was highly expressed in HTS-1 during the pistil-and-stamen-differentiating stage, at levels approximately 120 times greater than those in CM28TP. Further analysis indicated that TaWin1 was mainly expressed in HTS-1 PS, supporting its status as a candidate gene of hts. Thus, TaWin1 overexpression probably leads to the transformation of stamens into pistils in wheat. CONCLUSIONS: The results of this study provide a foundation for further research on stamen and pistil development, with implications for wheat-hybrid breeding programs.

Development of a high-density linkage map and mapping of the three-pistil gene (Pis1) in wheat using GBS markers.

BACKGROUND: The wheat mutant line three-pistil (TP) exhibits three pistils per floret. As TP normally has two or three seeds in each of the florets on the same spike, there is the possibility of increasing the number of grains per spike. Therefore, TP is a highly valuable mutant for breeding and for the study of floral development in wheat. To map the three-pistil gene (Pis1), genotyping-by-sequencing single-nucleotide polymorphism (GBS-SNP) data from an F2 mapping population (CM28 × CM28TP) was used to construct a genetic map that is of significant value. RESULTS: In the present study, a high-density genetic map of wheat containing 2917 GBS-SNP markers was constructed. Twenty-one linkage groups were resolved, with a total length of 2371.40 cM. The individual chromosomes range from 2.64 cM to 454.55 cM with an average marker density of 0.81 cM. The Pis1 gene was mapped using this high-resolution map, and two flanking SNP markers tightly linked to the gene, M70 and M71, were identified. The Pis1 is 3.00 cM from M70 and 1.10 cM from M71. In bread wheat genome, M70 and M71 were found to delimit a physical distance of 3.40 Mb, which encompasses 127 protein-coding genes. To validate the GBS-generated genotypic data and to eliminate missing marker data in the Pis1 region, five Kompetitive Allele-Specific PCR (KASP) assays were designed from corresponding GBS sequences, which harbor SNPs that surround Pis1. Three KASP-SNP markers, KM70, KM71, and KM75, were remapped to the Pis1 gene region. CONCLUSIONS: This work not only lays the foundation for the map-based cloning of Pis1 but can also serve as a valuable tool for studying marker-trait association of important traits and marker-assisted breeding in wheat.

Pistillody mutant reveals key insights into stamen and pistil development in wheat (Triticum aestivum L.).

BACKGROUND: The pistillody mutant wheat (Triticum aestivum L.) plant HTS-1 exhibits homeotic transformation of stamens into pistils or pistil-like structures. Unlike common wheat varieties, HTS-1 produces three to six pistils per floret, potentially increasing the yield. Thus, HTS-1 is highly valuable in the study of floral development in wheat. In this study, we conducted RNA sequencing of the transcriptomes of the pistillody stamen (PS) and the pistil (P) from HTS-1 plants, and the stamen (S) from the non-pistillody control variety Chinese Spring TP to gain insights into pistil and stamen development in wheat. RESULTS: Approximately 40 Gb of processed reads were obtained from PS, P, and S. De novo assembly yielded 121,210 putative unigenes, with a mean length of 695 bp. Among these high-quality unigenes, 59,199 (48.84%) had at least one significant match with an existing gene model. A total of 23, 263, and 553 differentially expressed genes were identified in PS vs. P, PS vs. S, and P vs. S, respectively, with differences in expression greater than five-fold. Among the differentially expressed genes, 206 were highly correlated with stamen and pistil development. These genes include WM27B, DL, YAB1, YABBY4, WM 5, CER 1, and WBLH1, which have been implicated in flower development. The expression patterns of 25 differentially expressed genes were confirmed through quantitative real-time reverse transcription PCR. CONCLUSIONS: Analysis of this transcriptome resource enabled us to characterize gene expression profiles, examine differential gene expression, and produce a candidate gene list related to wheat stamen and pistil development. This work is significant for the development of genomic resources for wheat, and provides important insights into the molecular mechanisms of wheat stamen and pistil development.

Over-expression of a plant-type phosphoenolpyruvate carboxylase derails Arabidopsis stamen formation.

We examined whether plant-type phosphoenolpyruvate carboxylase (PEPC) is involved in flower organ formation or not by over-expression in Arabidopsis. A wheat PEPC isogene Tappc3A, belonging to the ppc3 group, was targeted due to its preferential expression pattern in pistils and stamens. Transgenic Arabidopsis over-expressing Tappc3A exhibited irregular stamen formation, i.e., a lesser number of stamens per flower and shorter filaments in T2 and T3 generations. Irregular stamens were frequently observed in homozygous T4 lines, but no morphological change was observed in other floral organs. High-degree gene co-expression of Tappc3 isogenes with wheat SEEDSTICKs but not with other homeotic transcription factor genes for flower formation implicates that Tappc3 is under control by the class D genes of the ABCDE model to flower development. In addition, the conservation of CArG box sequences on the Tappc3 promoters supported the developmentally programmed gene expression of ppc3 in wheat flowering organs. Thus, this study provides the first experimental evidence for the critical regulation of plant-type PEPC for flower formation.

Characterization of the physiological parameters, effective components, and transcriptional profiles of Polygonum multiflorum Thunb. Under pH stress.

Polygonum multiflorum Thunb. is a traditional Chinese medicine with extensive distribution and robust adaptability, but comprehensive research on its acid and alkali resistance is presently lacking. This study aimed to analyze the effects of 5 months of continuous pH stress on the physiological and photosynthetic parameters of P. multiflorum, and the content of effective components. Results revealed that pH stress significantly influenced the normal growth, physiological functions, and photosynthetic indicators of P. multiflorum. At soil pH 4.5, the tubers of P. multiflorum exhibited the highest levels of 2,3,5,4'-tetrahydroxy stilbene-2-O-ß-d-glucoside (THSG) and total anthraquinones at 5.41% and 0.38%, respectively. However, increased soil pH significantly reduced the content of THSG and total anthraquinones. Reference-free transcriptome analysis was further conducted on P. multiflorum treated at pH 4.5 and 9.5, generating a total of 47,305 unigenes with an N50 of 2118 bp, of which 31,058 (65.65%) were annotated. Additionally, 2472 differentially expressed genes (DEGs) were identified. Among them, 17 DEGs associated with the biosynthesis of THSG and anthraquinones were screened. A comprehensive analysis of differential gene expression and effective component content demonstrated a significant positive correlation between the content of effective components and the 14 DEGs' expression but a negative correlation with soil pH. This study highlighted the influence of varying soil pH values on the effective component content of P. multiflorum. Specific acidic conditions proved beneficial for the synthesis and accumulation of THSG and total anthraquinones in P. multiflorum, thereby enhancing the quality of the medicinal material.

Host genetic variation and specialized metabolites from wheat leaves enriches for phyllosphere Pseudomonas spp. with enriched antibiotic resistomes.

Antibiotic resistance in plant-associated microbiomes poses significant risks for agricultural ecosystems and human health. Although accumulating evidence suggests a role for plant genotypes in shaping their microbiome, almost nothing is known about how the changes of plant genetic information affect the co-evolved plant microbiome carrying antibiotic resistance genes (ARGs). Here, we selected 16 wheat cultivars and experimentally explored the impact of host genetic variation on phyllosphere microbiome, ARGs, and metabolites. Our results demonstrated that host genetic variation significantly influenced the phyllosphere resistomes. Wheat genotypes exhibiting high phyllosphere ARGs were linked to elevated Pseudomonas populations, along with increased abundances of Pseudomonas aeruginosa biofilm formation genes. Further analysis of 350 Pseudomonas spp. genomes from diverse habitats at a global scale revealed that nearly all strains possess multiple ARGs, virulence factor genes (VFGs), and mobile genetic elements (MGEs) on their genomes, albeit with lower nucleotide diversity compared to other species. These findings suggested that the proliferation of Pseudomonas spp. in the phyllosphere significantly contributed to antibiotic resistance. We further observed direct links between the upregulated leaf metabolite DIMBOA-Glc, Pseudomonas spp., and enrichment of phyllosphere ARGs, which were corroborated by microcosm experiments demonstrating that DIMBOA-Glc significantly enhanced the relative abundance of Pseudomonas spp. Overall, alterations in leaf metabolites resulting from genetic variation throughout plant evolution may drive the development of highly specialized microbial communities capable of enriching phyllosphere ARGs. This study enhances our understanding of how plants actively shape microbial communities and clarifies the impact of host genetic variation on the plant resistomes.

Cloning and characterization of endo-ß-1,4-glucanase genes in the common wheat line three pistils.

In this work, we report the cloning and characterization of endo-ß-1,4-glucanase (EGase) genes (TaEG) in the common wheat line three pistils. Three TaEG homoeologous genes (TaEG-4A, TaEG-4B and TaEG-4D) were isolated and found to be located on chromosomes 4AL, 4BS and 4DS, respectively. The three genes showed high conservation of their coding nucleotide sequences and 3 untranslated region. The putative TaEG protein had a molecular mass of 69 kDa, a theoretical pI of 9.39 and a transmembrane domain of 74-96 amino acids in the N-terminus that anchored the protein to the membrane. The genome sequences of TaEG-4A, TaEG-4B and TaEG-4D contained six exons and five introns. All of the introns, except for intron IV, varied in length and sequence composition. Phylogenetic analysis revealed that TaEG was most closely related to rice (Oryza sativa) OsGLU1. The TaEG transcript levels increased significantly during the subsidiary pistil primordium differentiation phase (spike size ∼7-10 mm) in Chuanmai 28 TP (CM28TP). These data provide a basis for future research into the function of TaEG and offer insights into the molecular mechanism of the three pistils mutation in wheat.

The Original Form of C4-Photosynthetic Phosphoenolpyruvate Carboxylase Is Retained in Pooids but Lost in Rice.

Poaceae is the most prominent monocot family that contains the primary cereal crops wheat, rice, and maize. These cereal species exhibit physiological diversity, such as different photosynthetic systems and environmental stress tolerance. Phosphoenolpyruvate carboxylase (PEPC) in Poaceae is encoded by a small multigene family and plays a central role in C4-photosynthesis and dicarboxylic acid metabolism. Here, to better understand the molecular basis of the cereal species diversity, we analyzed the PEPC gene family in wheat together with other grass species. We could designate seven plant-type and one bacterial-type grass PEPC groups, ppc1a, ppc1b, ppc2a, ppc2b, ppc3, ppc4, ppcC4, and ppc-b, respectively, among which ppc1b is an uncharacterized type of PEPC. Evolutionary inference revealed that these PEPCs were derived from five types of ancient PEPCs (ppc1, ppc2, ppc3, ppc4, and ppc-b) in three chromosomal blocks of the ancestral Poaceae genome. C4-photosynthetic PEPC (ppcC4 ) had evolved from ppc1b, which seemed to be arisen by a chromosomal duplication event. We observed that ppc1b was lost in many Oryza species but preserved in Pooideae after natural selection. In silico analysis of cereal RNA-Seq data highlighted the preferential expression of ppc1b in upper ground organs, selective up-regulation of ppc1b under osmotic stress conditions, and nitrogen response of ppc1b. Characterization of wheat ppc1b showed high levels of gene expression in young leaves, transcriptional responses under nitrogen and abiotic stress, and the presence of a Dof1 binding site, similar to ppcC4 in maize. Our results indicate the evolving status of Poaceae PEPCs and suggest the functional association of ppc1-derivatives with adaptation to environmental changes.

Co-expression network analysis of genes and networks associated with wheat pistillody.

Crop male sterility has great value in theoretical research and breeding application. HTS-1, whose stamens transformed into pistils or pistil-like structures, is an important male sterility material selecting from Chinese Spring three-pistil (CSTP) wheat. However the molecular mechanism of pistillody development in HTS-1 remains a mystery. RNA-seq data of 11 wheat tissues were obtained from the National Center for Biotechnology Information (NCBI), including the stamens of CSTP and the pistils and pistillodic stamen of HTS-1. The Salmon program was utilized to quantify the gene expression levels of the 11 wheat tissues; and gene quantification results were normalized by transcripts per million (TPM). In total, 58,576 genes were used to construct block-wise network by co-expression networks analysis (WGCNA) R package. We obtained all of modules significantly associated with the 11 wheat tissues. AgriGO V2.0 was used to do Gene Ontology (GO) enrichment analysis; and genes and transcription factors (TFs) in these significant modules about wheat pistillody development were identified from GO enrichment results. Basic local alignment search tool (BLAST) was used to align HTS-1 proteins with the published pistillody-related proteins and TFs. Genes about wheat pistillody development were analyzed and validated by qRT-PCR. The MEturquoise, MEsaddlebrown, MEplum, MEcoral1, MElightsteelblue1, and MEdarkslateblue modules were significantly corelated to pistillodic stamen (correlation p < 0.05). Moreover, 206 genes related to carpel development (GO:0048440) or gynoecium development (GO:0048467) were identified only in the MEturquoise module by Gene Ontology (GO) analysis, and 42 of 206 genes were hub genes in MEturquoise module. qRT-PCR results showed that 38 of the 42 hub genes had highly expressed in pistils and pistillodic stamens than in stamens. A total of 15 pistillody development-related proteins were validated by BLAST. Transcription factors (TFs) were also analyzed in the MEturquoise module, and 618 TFs were identified. In total, 56 TFs from 11 families were considered to regulate the development of pistillodic stamen. The co-expression network showed that six of HB and three of BES1 genes were identified in 42 hub genes. This indicated that TFs played important roles in wheat pistillody development. In addition, there were 11 of ethylene-related genes connected with TFs or hub genes, suggesting the important roles of ethylene-related genes in pistillody development. These results provide important insights into the molecular interactions underlying pistillody development.

Combined effects of liquid digestate recirculation and biochar on methane yield, enzyme activity, and microbial community during semi-continuous anaerobic digestion.

The combined effects of liquid digestate recirculation (LDR) and biochar on methanogenesis and microbial communities were studied in semi-continuous anaerobic reactors fed with wheat straw and swine manure. The tolerated organic loading rate (OLR) was expanded from 5 g- volatile solids (VS)∙L-1∙d-1 in the control to higher than 6 g-VS∙L-1∙d-1 in the LDR. At the OLR of 5.0 g-VS∙L-1∙d-1, average special methane yield in LDR with biochar was 0.234 L∙g-VS-1, which was 5.4 % higher than that of the LDR alone. Moreover, enzyme activity and microbial community analysis indicated that LDR with biochar enhanced the processes of hydrolysis and methanogenesis, and balanced the pathway between hydrogenotrophic and acetoclastic methanogenesis. The co-application of LDR and biochar synergistically enhanced the degradation pathways of substrates and the loading shock resistance of anaerobic digestion system. This study could offer strategies for developing sustainable applications of full and continuous LDR in industrial biogas projects.

Enhanced brassinosteroid signaling via the overexpression of SlBRI1 positively regulates the chilling stress tolerance of tomato.

Brassinosteroids (BRs) regulate plant development and response to stress. BRASSINOSTEROID INSENSITIVE 1 (BRI1) is a BR receptor that activates BR signaling. Although the function of the tomato BR receptor SlBRI1 in regulating growth and drought resistance has been examined, that of SlBRI1 in cold tolerance is unclear. This study indicated that the expression of SlBRI1 in tomato was rapidly induced and reached its highest level at 3 h under chilling treatment and then decreased. The overexpression of SlBRI1 displayed low relative electrolyte leakage, malondialdehyde content, and reactive oxygen species (ROS) accumulation under chilling stress. The proline content and activities of superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT) in SlBRI1OE plants were higher than those in the wild-type (WT) plants after chilling stress. The transcript abundances of five cold-responsive genes were higher in SlBRI1OE plants than in WT plants during chilling stress. RNA sequence analysis showed that the expression of the majority of genes encoding photosystem I and II were downregulated. The degree of suppression in SlBRI1OE plants was weaker than that in WT plants. Additionally, the Pn and Fv/Fm of SlBRI1OE plants were significantly higher than those of WT plants under chilling stress. We identified several genes encoding key enzymes in BRs; indole acetic acid (IAA), gibberellin (GA), and abscisic acid (ABA) biosynthesis or signaling were upregulated or downregulated during chilling stress. Chilling stress decreased the BRs and GA3 content, and increased IAA and ABA content. The contents were lower or higher in SlBRI1OE than in WT plants. Furthermore, chilling stress regulated the expression levels of 43 transcription factors. The expression of seven cold-regulated protein genes was higher or lower in SlBRI1OE plants than in WT plants under chilling stress. These results suggest that SlBRI1 positively regulates chilling tolerance mainly through ICE1-CBF-COR pathway in tomato.

RicetissueTFDB: A Genome-Wide Identification of Tissue-Specific Transcription Factors in Rice.

Transcription factors (TFs) regulate plant gene expression in different tissues. To investigate TF genes in rice ( L.), a genome-wide TF identification was conducted with the japonica rice genome. This study identified 3078 putative TFs in 59 families. The TF number of the top 10 TF families accounted for 58% of the 3078 rice TFs. The three largest TF families were the myeloblastosis (MYB) superfamily, basic helix-loop-helix (bHLH), and far-red-impaired response (FAR1), which contained 413, 228, and 210 TF members, respectively. The expression profiles of the 3078 TF genes were surveyed with the RNA sequencing (RNA-seq) data of 13 rice tissue types. Based on these expression profiles, we validated 1087 TFs expressed in 13 rice tissue types, which accounted for 35.32% of the 3078 putative TFs. We further analyzed the tissue-specific TFs in rice. In total, 28, 14, 14, 10, 9, 5, 5, 4, 3, 3, 2, 11, and 1 tissue-specific TF sequences were identified in the dry seed, pistil, spikelet, aleurone, anther, ovules, embryo 25 d after pollination (DAP), seed 5 DAP, root, leaf, seed 10 DAP, shoot, and endosperm 25 DAP, respectively. Moreover, we constructed RicetissueTFDB (), a comprehensive and public rice TF database that integrates tissue expression characters, genomic location, and Gene Ontology (GO) terms for each TF. The RicetissueTFDB database will facilitate the identification of target TFs and the functional studies about rice TFs.