Temporospatial pattern of flavonoid metabolites and potential regulatory pathway of PbMYB211-coordinated kaempferol-3-O-rhamnoside biosynthesis in Phoebe bournei.

Phoebe is a well-known timber tree species that contains abundant metabolites characterized by flavonoids that are widely used in the pharmaceutical industry. Nevertheless, temporospatial flavonoid metabolism variations substantially impact the Phoebe industry. Thus, a metabolomics analysis was carried out and identified 465 metabolites (102 flavonoids) in P. bournei, revealing distinct distribution patterns among five studied organs, and most of the flavonoids were dominant in the leaves. Furthermore, three kaempferol glycoside derivatives were significantly accumulated in the leaves and showed higher contents in young leaves than in mature leaves and differences between spring and autumn. For instance, greater accumulation of kaempferol-7-O-rhamnoside was detected in spring, whereas higher contents of kaempferol-3-O-arabinofuranoside and kaempferol-3-O-rhamnoside were found in autumn. Integrated metabolomics and transcriptomics identified 20 transcription factors (TFs) and 12 structural genes that participate in kaempferol derivative synthesis and elucidated a potential regulatory mechanism in P. bournei. Of the identified genes, PbMYB211 might contribute significantly to the kaempferol-3-O-rhamnoside content by regulating the target structural gene PbUGT139, as revealed by transient overexpression analysis. Overall, this study illuminated the temporospatial accumulation of flavonoids among different organs, seasons, and developmental stages in P. bournei and elucidated a potential regulatory pathway of kaempferol-3-O-rhamnoside. The results provide important insights into harvest techniques and a theoretical basis for the comprehensive utilization of P. bournei.

Melatonin-mediated development and abiotic stress tolerance in plants.

Melatonin is a multifunctional molecule that has been widely discovered in most plants. An increasing number of studies have shown that melatonin plays essential roles in plant growth and stress tolerance. It has been extensively applied to alleviate the harmful effects of abiotic stresses. In view of its role in regulating aspects of plant growth and development, we ponder and summarize the scientific discoveries about seed germination, root development, flowering, fruit maturation, and senescence. Under abiotic and biotic stresses, melatonin brings together many pathways to increase access to treatments for the symptoms of plants and to counteract the negative effects. It has the capacity to tackle regulation of the redox, plant hormone networks, and endogenous melatonin. Furthermore, the expression levels of several genes and the contents of diverse secondary metabolites, such as polyphenols, terpenoids, and alkaloids, were significantly altered. In this review, we intend to examine the actions of melatonin in plants from a broader perspective, explore the range of its physiological functions, and analyze the relationship between melatonin and other metabolites and metabolic pathways.

Cyanidin-3-O-glucoside Contributes to Leaf Color Change by Regulating Two bHLH Transcription Factors in Phoebe bournei.

Anthocyanins produce different-colored pigments in plant organs, which provide ornamental value. Thus, this study was conducted to understand the mechanism of anthocyanin synthesis in ornamental plants. Phoebe bournei, a Chinese specialty tree, has high ornamental and economic value due to its rich leaf color and diverse metabolic products. Here, the metabolic data and gene expression of red P. bournei leaves at the three developmental stages were evaluated to elucidate the color-production mechanism in the red-leaved P. bournei species. First, metabolomic analysis identified 34 anthocyanin metabolites showing high levels of cyanidin-3-O-glucoside (cya-3-O-glu) in the S1 stage, which may suggest that it is a characteristic metabolite associated with the red coloration of the leaves. Second, transcriptome analysis showed that 94 structural genes were involved in anthocyanin biosynthesis, especially flavanone 3'-hydroxy-lase (PbF3'H), and were significantly correlated with the cya-3-O-glu level. Third, K-means clustering analysis and phylogenetic analyses identified PbbHLH1 and PbbHLH2, which shared the same expression pattern as most structural genes, indicating that these two PbbHLH genes may be regulators of anthocyanin biosynthesis in P. bournei. Finally, overexpression of PbbHLH1 and PbbHLH2 in Nicotiana tabacum leaves triggered anthocyanin accumulation. These findings provide a basis for cultivating P. bournei varieties that have high ornamental value.

Genome-Wide Identification and Expression Analysis of Calmodulin and Calmodulin-like Genes, Revealing CaM3 and CML13 Participating in Drought Stress in Phoebe bournei.

Calmodulin (CaM) and calmodulin-like (CML) proteins are major Ca2+ sensors involved in the regulation of plant development and stress responses by converting Ca2+ signals into appropriate cellular responses. However, characterization and expression analyses of CaM/CML genes in the precious species, Phoebe bournei, remain limited. In this study, five PbCaM and sixty PbCML genes were identified that only had EF-hand motifs with no other functional domains. The phylogenetic tree was clustered into 11 subgroups, including a unique clade of PbCaMs. The PbCaMs were intron-rich with four EF-hand motifs, whereas PbCMLs had two to four EF-hands and were mostly intronless. PbCaMs/CMLs were unevenly distributed across the 12 chromosomes of P. bournei and underwent purifying selection. Fragment duplication was the main driving force for the evolution of the PbCaM/CML gene family. Cis-acting element analysis indicated that PbCaMs/CMLs might be related to hormones, growth and development, and stress response. Expression analysis showed that PbCaMs were generally highly expressed in five different tissues and under drought stress, whereas PbCMLs showed specific expression patterns. The expression levels of 11 candidate PbCaMs/CMLs were responsive to ABA and MeJA, suggesting that these genes might act through multiple signaling pathways. The overexpression of PbCaM3/CML13 genes significantly increased the tolerance of yeast cells to drought stress. The identification and characterization of the CaM/CML gene family in P. bournei laid the foundation for future functional studies of these genes.

Co-expression network analysis reveals PbTGA4 and PbAPRR2 as core transcription factors of drought response in an important timber species Phoebe bournei.

Phoebe bournei is one of the main afforestation tree species in subtropical regions of China and is famous for its timber. Its distribution and growth are significantly impaired by water conditions. Thus, it is essential to understand the mechanism of the stress response in P. bournei. Here, we analyzed the phenotypic changes and transcriptomic rearrangement in the leaves and roots of P. bournei seedlings grown for 0 h, 1 h, 24 h, and 72 h under simulated drought conditions (10% PEG 6000). The results showed that drought stress inhibited plant photosynthesis and increased oxidoreductase activity and abscisic acid (ABA) accumulation. Spatio-temporal transcriptomic analysis identified 2836 and 3704 differentially expressed genes (DEGs) in leaves and roots, respectively. The responsive genes in different organs presented various expression profiles at different times. Gene co-expression network analysis identified two core transcription factors, TGA4 and APRR2, from two modules that showed a strong positive correlation with ABA accumulation. Our study investigated the different responses of aboveground and belowground organs of P. bournei to drought stress and provides critical information for improving the drought resistance of this timber species.

Comparative physiological analyses and the genetic basis reveal heat stress responses mechanism among different Betula luminifera populations.

Betula luminifera is a subtropical fast-growing timber species with high economic value. However, along with global warming, heat stress become one of the main environmental variables that limit the productivity of B. luminifera, and the response of diverse geographic populations to high temperatures is still unclear. In order to offer a comprehensive understanding of the behavior of B. luminifera under heat stress, the physiological responses of six B. luminifera populations (across the core distribution area) were described in this work in an integrated viewpoint. The results showed that a multi-level physiological regulatory network may exist in B. luminifera, the first response was the activity of resistant enzymes [e.g., peroxidase (POD)] at a preliminary stage of 2 h heat stress, and then the proline (osmoregulation substance) content began to increase after 24 h of continuous high-temperature treatment. In addition, photosynthesis was stronlgly affected by heat stress, and the net photosynthetic rate (Pn ) showed a downward trend under heat treatment in all six B. luminifera populations. Interestingly, although the physiological change patterns of the six B. luminifera populations were relatively consistent for the same parameter, there were obvious differences among different populations. Comprehensive analysis revealed that the physiological response of Rongshui (RS) was the most stable, and this was the representative B. luminifera population. Illumina RNA-seq analysis was applied to reveal the specific biological process of B. luminifera under heat stress using the RS population, and a total of 116,484 unigenes were obtained. The differentially expressed genes (DEGs) between different time periods under heat stress were enriched in 34 KEGG pathways, and the limonene and pinene degradation pathway was commonly enriched in all pairwise comparisons. Moreover, transcription factors including bHLH (basic helix-loop-helix), MYB, WRKY, and NAC (NAM, ATAF1/2, and CUC2) were identified. In this study, the physiological response and tolerance mechanisms of B. luminifera under high temperature stress were revealed, which can conducive to the basis of B. luminifera selection and resistance assessment for cultivation and breeding.

Identification of Nitrogen Starvation-Responsive miRNAs to Reveal the miRNA-Mediated Regulatory Network in Betula luminifera.

Because of the immobility, plants encounter a series of stresses, such as varied nutrient concentrations in soil, which regulate plant growth, development, and phase transitions. Nitrogen (N) is one of the most limiting factors for plants, which was exemplified by the fact that low nitrogen (LN) has a great adverse effect on plant growth and development. In the present study, we explored the potential role of microRNAs (miRNAs) in response to LN stress in Betula luminifera. We identified 198 miRNAs using sRNA sequencing, including 155 known and 43 novel miRNAs. Among them, 98 known miRNAs and 31 novel miRNAs were differentially expressed after 0.5 h or 24 h of LN stress. Based on degradome data, 122 differential expressed miRNAs (DEmiRNAs) including 102 known miRNAs and 20 novel miRNAs targeted 203 genes, comprising 321 miRNA-target pairs. A big proportion of target genes were transcription factors and functional proteins, and most of the Gene Ontology terms were enriched in biological processes; moreover, one Kyoto Encyclopedia of Genes and Genomes term "ascorbate and aldarate metabolism" was significantly enriched. The expression patterns of six miRNAs and their corresponding target genes under LN stress were monitored. According to the potential function for targets of DEmiRNAs, a proposed regulatory network mediated by miRNA-target pairs under LN stress in B. luminifera was constructed. Taken together, these findings provide useful information to elucidate miRNA functions and establish a framework for exploring N signaling networks mediated by miRNAs in B. luminifera. It may provide new insights into the genetic engineering of the high use efficiency of N in forestry trees.

Assessing the current genetic structure of 21 remnant populations and predicting the impacts of climate change on the geographic distribution of Phoebe sheareri in southern China.

Phoebe sheareri is a valuable tree species known as "Golden Nanmu" and is one of the most important protected tree species in China. However, natural populations are decreasing because of climate change and anthropogenic factors. To evaluate the genetic diversity and structure of remnant populations and the impacts of climate change on the distribution of potential suitable habitats, we conducted a field investigation and sampled 21 P. sheareri natural populations to evaluate their genetic diversity and structure using simple sequence repeat (SSR) molecular markers. Then, we predicted the distribution of suitable P. sheareri habitats across China under future scenarios (RCP 2.6 and RCP 8.5) and periods (2050 and 2070) using multivariate modeling methods-the MaxEnt model. The results showed a medium level of genetic diversity and low inbreeding in the 21 P. sheareri natural populations, and genetic differentiation among populations was significant, with 21.2 % genetic variation among populations. The remnant populations of P. sheareri were grouped into four genetic clusters based on genetic structure; five environmental variables involving four temperature variables and precipitation seasonality (Bio12) might determine the distribution of P. sheareri populations. In the future, the suitable habitats of P. sheareri are manifested as northward migration, and the highly suitable habitats are expected to increase. Our results highlight the importance of conservation units in situ, giving priority to populations with higher genetic diversity (e.g., TMS, FJS, and THY populations); sampling strategies for ex situ conservation, breeding and reforestation should consider climate change, especially Bio1 (annual mean temperature) and Bio12 (annual precipitation). Overall, this study may provide useful genetic information for strategies for the protection, management, and utilization of P. sheareri.

The chromosome-scale genome of Phoebe bournei reveals contrasting fates of terpene synthase (TPS)-a and TPS-b subfamilies.

Terpenoids, including aromatic volatile monoterpenoids and sesquiterpenoids, function in defense against pathogens and herbivores. Phoebe trees are remarkable for their scented wood and decay resistance. Unlike other Lauraceae species investigated to date, Phoebe species predominantly accumulate sesquiterpenoids instead of monoterpenoids. Limited genomic data restrict the elucidation of terpenoid variation and functions. Here, we present a chromosome-scale genome assembly of a Lauraceae tree, Phoebe bournei, and identify 72 full-length terpene synthase (TPS) genes. Genome-level comparison shows pervasive lineage-specific duplication and contraction of TPS subfamilies, which have contributed to the extreme terpenoid variation within Lauraceae species. Although the TPS-a and TPS-b subfamilies were both expanded via tandem duplication in P. bournei, more TPS-a copies were retained and constitutively expressed, whereas more TPS-b copies were lost. The TPS-a genes on chromosome 8 functionally diverged to synthesize eight highly accumulated sesquiterpenes in P. bournei. The essential oil of P. bournei and its main component, ß-caryophyllene, exhibited antifungal activities against the three most widespread canker pathogens of trees. The TPS-a and TPS-b subfamilies have experienced contrasting fates over the evolution of P. bournei. The abundant sesquiterpenoids produced by TPS-a proteins contribute to the excellent pathogen resistance of P. bournei trees. Overall, this study sheds light on the evolution and adaptation of terpenoids in Lauraceae and provides valuable resources for boosting plant immunity against pathogens in various trees and crops.

Full-Length Transcriptome Sequencing and Comparative Transcriptomic Analyses Provide Comprehensive Insight Into Molecular Mechanisms of Cellulose and Lignin Biosynthesis in Cunninghamia lanceolata.

Cunninghamia lanceolata is an essential timber species that provide 20%-30% raw materials for China's timber industry. Although a few transcriptomes have been published in C. lanceolata, full-length mRNA transcripts and regulatory mechanisms behind the cellulose and lignin biosynthesis have not been thoroughly investigated. Here, PacBio Iso-seq and RNA-seq analyses were adapted to identify the full-length and differentially expressed transcripts along a developmental gradient from apex to base of C. lanceolata shoots. A total of 48,846 high-quality full-length transcripts were obtained, of which 88.0% are completed transcriptome based on benchmarking universal single-copy orthologs (BUSCO) assessment. Along stem developmental gradient, 18,714 differentially expressed genes (DEGs) were detected. Further, 28 and 125 DEGs were identified as enzyme-coding genes of cellulose and lignin biosynthesis, respectively. Moreover, 57 transcription factors (TFs), including MYB and NAC, were identified to be involved in the regulatory network of cellulose and lignin biosynthesis through weighted gene co-expression network analysis (WGCNA). These TFs are composed of a comparable regulatory network of secondary cell wall formation in angiosperms, revealing a similar mechanism may exist in gymnosperms. Further, through qRT-PCR, we also investigated eight specific TFs involved in compression wood formation. Our findings provide a comprehensive and valuable source for molecular genetics breeding of C. lanceolata and will be beneficial for molecular-assisted selection.

Rice LEAFY COTYLEDON1 Hinders Embryo Greening During the Seed Development.

LEAFY COTYLEDON1 (LEC1) is the central regulator of seed development in Arabidopsis, while its function in monocots is largely elusive. We generated Oslec1 mutants using CRISPR/Cas9 technology. Oslec1 mutant seeds lost desiccation tolerance and triggered embryo greening at the early development stage. Transcriptome analysis demonstrated that Oslec1 mutation altered diverse hormonal pathways and stress response in seed maturation, and promoted a series of photosynthesis-related genes. Further, genome-wide identification of OsLEC1-binding sites demonstrated that OsLEC1 bound to genes involved in photosynthesis, photomorphogenesis, as well as abscisic acid (ABA) and gibberellin (GA) pathways, involved in seed maturation. We illustrated an OsLEC1-regulating gene network during seed development, including the interconnection between photosynthesis and ABA/GA biosynthesis/signaling. Our findings suggested that OsLEC1 acts as not only a central regulator of seed maturation but also an inhibitor of embryo greening during rice seed development. This study would provide new understanding for the OsLEC1 regulatory mechanisms on photosynthesis in the monocot seed development.

Full-length transcriptomic identification of R2R3-MYB family genes related to secondary cell wall development in Cunninghamia lanceolata (Chinese fir).

BACKGROUND: R2R3-MYB is a class of transcription factor crucial in regulating secondary cell wall development during wood formation. The regulation of wood formation in gymnosperm has been understudied due to its large genome size. Using Single-Molecule Real-Time sequencing, we obtained full-length transcriptomic libraries from the developmental stem of Cunninghamia lanceolata, a perennial conifer known as Chinese fir. The R2R3-MYB of C. lanceolata (hereafter named as ClMYB) associated with secondary wall development were identified based on phylogenetic analysis, expression studies and functional study on transgenic line. RESULTS: The evolutionary relationship of 52 ClMYBs with those from Arabidopsis thaliana, Eucalyptus grandis, Populus trichocarpa, Oryza sativa, two gymnosperm species, Pinus taeda, and Picea glauca were established by neighbour-joining phylogenetic analysis. A large number of ClMYBs resided in the woody-expanded subgroups that predominated with the members from woody dicots. In contrast, the woody-preferential subgroup strictly carrying the members of woody dicots contained only one candidate. The results suggest that the woody-expanded subgroup emerges before the gymnosperm/angiosperm split, while most of the woody-preferential subgroups are likely lineage-specific to woody dicots. Nine candidates shared the same subgroups with the A. thaliana orthologs, with known function in regulating secondary wall development. Gene expression analysis inferred that ClMYB1/2/3/4/5/26/27/49/51 might participate in secondary wall development, among which ClMYB1/2/5/26/27/49 were significantly upregulated in the highly lignified compression wood region, reinforcing their regulatory role associated with secondary wall development. ClMYB1 was experimentally proven a transcriptional activator that localised in the nucleus. The overexpression of ClMYB1 in Nicotiana benthamiana resulted in an increased lignin deposition in the stems. The members of subgroup S4, ClMYB3/4/5 shared the ERF-associated amphiphilic repression motif with AtMYB4, which is known to repress the metabolism of phenylpropanoid derived compounds. They also carried a core motif specific to gymnosperm lineage, suggesting divergence of the regulatory process compared to the angiosperms. CONCLUSIONS: This work will enrich the collection of full-length gymnosperm-specific R2R3-MYBs related to stem development and contribute to understanding their evolutionary relationship with angiosperm species.

Comprehensive Analysis of Five Phyllostachys edulis SQUA-like Genes and Their Potential Functions in Flower Development.

Bamboo is one of the most important non-timber forest resources worldwide. It has considerable economic value and unique flowering characteristics. The long juvenile phase in bamboo and unpredictable flowering time limit breeding and genetic improvement and seriously affect the productivity and application of bamboo forests. Members of SQUA-like subfamily genes play an essential role in controlling flowering time and floral organ identity. A comprehensive study was conducted to explain the functions of five SQUA-like subfamily genes in Phyllostachys edulis. Expression analysis revealed that all PeSQUAs have higher transcript levels in the reproductive period than in the juvenile phase. However, PeSQUAs showed divergent expression patterns during inflorescence development. The protein-protein interaction (PPI) patterns among PeSQUAs and other MADS-box members were analyzed by yeast two-hybrid (Y2H) experiments. Consistent with amino acid sequence similarity and phylogenetic analysis, the PPI patterns clustered into two groups. PeMADS2, 13, and 41 interacted with multiple PeMADS proteins, whereas PeMADS3 and 28 hardly interacted with other proteins. Based on our results, PeSQUA might possess different functions by forming protein complexes with other MADS-box proteins at different flowering stages. Furthermore, we chose PeMADS2 for functional analysis. Ectopic expression of PeMADS2 in Arabidopsis and rice caused early flowering, and abnormal phenotype was observed in transgenic Arabidopsis lines. RNA-seq analysis indicated that PeMADS2 integrated multiple pathways regulating floral transition to trigger early flowering time in rice. This function might be due to the interaction between PeMADS2 and homologous in rice. Therefore, we concluded that the five SQUA-like genes showed functional conservation and divergence based on sequence differences and were involved in floral transitions by forming protein complexes in P. edulis. The MADS-box protein complex model obtained in the current study will provide crucial insights into the molecular mechanisms of bamboo's unique flowering characteristics.

Profiling of MicroRNAs and Their Targets in Roots and Shoots Reveals a Potential MiRNA-Mediated Interaction Network in Response to Phosphate Deficiency in the Forestry Tree Betula luminifera.

Inorganic phosphate (Pi) is often lacking in natural and agro-climatic environments, which impedes the growth of economically important woody species. Plants have developed strategies to cope with low Pi (LP) availability. MicroRNAs (miRNAs) play important roles in responses to abiotic stresses, including nutrition stress, by regulating target gene expression. However, the miRNA-mediated regulation of these adaptive responses and their underlying coordinating signals are still poorly understood in forestry trees such as Betula luminifera. Transcriptomic libraries, small RNA (sRNA) libraries, and a mixed degradome cDNA library of B. luminifera roots and shoots treated under LP and normal conditions (CK) were constructed and sequenced using next-generation deep sequencing. A comprehensive B. luminifera transcriptome derived from its roots and shoots was constructed, and a total of 76,899 unigenes were generated. Analysis of the transcriptome identified 8,095 and 5,584 differentially expressed genes in roots and shoots, respectively, under LP conditions. sRNA sequencing analyses indicated that 66 and 60 miRNAs were differentially expressed in roots and shoots, respectively, under LP conditions. A total of 109 and 112 miRNA-target pairs were further validated in the roots and shoots, respectively, using degradome sequencing. Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis of differential miRNA targets indicated that the "ascorbate and aldarate metabolism" pathway responded to LP, suggesting miRNA-target pairs might participating in the removing of reactive oxidative species under LP stress. Moreover, a putative network of miRNA-target interactions involved in responses to LP stress in B. luminifera is proposed. Taken together, these findings provide useful information to decipher miRNA functions and establish a framework for exploring P signaling networks regulated by miRNAs in B. luminifera and other woody plants. It may provide new insights into the genetic engineering of high use efficiency of Pi in forestry trees.

Simultaneous detection of zearalenone, citrinin, and ochratoxin A in pepper by capillary zone electrophoresis.

In the present study, a simple and fast method for simultaneous detection of zearalenone, citrinin, and ochratoxin A utilising capillary zone electrophoresis with an ultraviolet detector was developed. The optimised approach was validated and applied using pepper samples. The proposed method yielded satisfactory linearity between the signal and the mycotoxin concentration in the range of 1.5-150 µg/kg for zearalenone, 4.5-150 µg/kg for citrinin, and 0.8-150 µg/kg for ochratoxin A. The limits of detection for these mycotoxins ranged from 0.3 to 1.5 µg/kg. The corresponding intra- and inter-day precisions were less than 3.5 % and 4.1 %, respectively. Moreover, the matrix effect was also assessed and the result was compared using the capillary zone electrophoresis and high-performance liquid chromatography methods. The developed approach could be used for simultaneous detection of zearalenone, citrinin, and ochratoxin A in pepper.

Colchicine-Induced Polyploidy in Rhododendron fortunei Lindl.

Polyploidy in Rhododendron fortunei has great potential to improve its horticultural and commercial value, and to also meet market demands. In this study, a feasible method for polyploid induction in R. fortunei via colchicine treatment was established, and the obtained polyploid plants were identified and characterized. As a result, the stem bases of tissue-cultured plantlets treated with 0.1% colchicine for 24 h showed the highest polyploid induction with a rate of 36.67%. By flow cytometric analysis, 69 tetraploids and 29 octoploids were identified in the regenerated plants that were examined. Phenotypic analysis indicated that the leaves of tetraploid and octoploid plants were smaller, rounder and thicker with more abundant and longer epidermal hairs than those of diploids. Furthermore, the stomata of polyploids were larger and sparser than those of diploids. An increase in chlorophyll content was also detected in polyploids, which resulted in darker green leaves. In conclusion, our study established an effective method to induce polyploidy in R. fortunei, which could be used to develop new genetic resources for breeding R. fortunei and other Rhododendron species in the future.

Genome survey of Chinese fir (Cunninghamia lanceolata): Identification of genomic SSRs and demonstration of their utility in genetic diversity analysis.

Chinese fir (Cunninghamia lanceolata) is an important coniferous species that accounts for 20-30% of the total commercial timber production in China. Though traditional breeding of Chinese fir has achieved remarkable success, molecular-assisted breeding has made little progress due to limited availability of genomic information. In this study, a survey of Chinese fir genome was performed using the Illumina HiSeq Xten sequencing platform. K-mer analysis indicated that Chinese fir has a large genome of approximately 11.6 Gb with 74.89% repetitive elements and is highly heterozygous. Meanwhile, its genome size was estimated to be 13.2 Gb using flow cytometry. A total of 778.02 Gb clean reads were assembled into 10,982,272 scaffolds with an N50 of 1.57 kb. In total, 362,193 SSR loci were detected with a frequency of 13.18 kb. Dinucleotide repeats were the most abundant (up to 73.6% of the total SSRs), followed by trinucleotide and tetranucleotide repeats. Forty-six polymorphic pairs were developed, and 298 alleles were successfully amplified from 199 Chinese fir clones. The average PIC value was 0.53, indicating that the identified genomic SSR (gSSR) markers have a high degree of polymorphism. In addition, these breeding resources were divided into three groups, and a limited gene flow existed among these inferred groups.

Genome-wide identification of MADS-box family genes in moso bamboo (Phyllostachys edulis) and a functional analysis of PeMADS5 in flowering.

BACKGROUND: MADS-box genes encode a large family of transcription factors that play significant roles in plant growth and development. Bamboo is an important non-timber forest product worldwide, but previous studies on the moso bamboo (Phyllostachys edulis) MADS-box gene family were not accurate nor sufficiently detailed. RESULTS: Here, a complete genome-wide identification and characterization of the MADS-box genes in moso bamboo was conducted. There was an unusual lack of type-I MADS-box genes in the bamboo genome database ( http://202.127.18.221/bamboo/index.php ), and some of the PeMADS sequences are fragmented and/or inaccurate. We performed several bioinformatics techniques to obtain more precise sequences using transcriptome assembly. In total, 42 MADS-box genes, including six new type-I MADS-box genes, were identified in bamboo, and their structures, phylogenetic relationships, predicted conserved motifs and promoter cis-elements were systematically investigated. An expression analysis of the bamboo MADS-box genes in floral organs and leaves revealed that several key members are involved in bamboo inflorescence development, like their orthologous genes in Oryza. The ectopic overexpression of one MADS-box gene, PeMADS5, in Arabidopsis triggered an earlier flowering time and the development of an aberrant flower phenotype, suggesting that PeMADS5 acts as a floral activator and is involved in bamboo flowering. CONCLUSION: We produced the most comprehensive information on MADS-box genes in moso bamboo. Additionally, a critical PeMADS gene (PeMADS5) responsible for the transition from vegetative to reproductive growth was identified and shown to be related to bamboo floral development.

RNA-Seq analysis of differential gene expression in Betula luminifera xylem during the early stages of tension wood formation.

BACKGROUND: Betula luminifera H. Winkler, which is widely distributed in southern China, is an economically important broadleaf tree species. However, little genomic information of B. luminifera is available, and little is known about the molecular mechanisms of wood formation in this species. Meanwhile, few efforts have focused on investigating the early transcriptional changes during tension wood formation in woody plants. RESULTS: A reference transcriptome dataset was first generated containing 45,700 Unigenes, and 35,135 (76.9%) Unigenes were annotated by a BLAST similarity search against four public databases. Then, based on an anatomical investigation, the global gene expression changes during the early stages of tension wood formation were analyzed. Gene expression profiling showed that a total of 13,273 Unigenes were differentially regulated during the early stages of tension wood formation. Most genes involved in cellulose and lignin biosynthesis were highlighted to reveal their biological importance in tension wood formation. In addition, the transcription levels of many genes involved in the auxin response pathway were significantly changed during the early stages of tension wood formation. Furthermore, 18 TFs co-expressed with key enzymes of cellulose synthesis were identified. CONCLUSIONS: Our results revealed the transcriptional changes associated with TW formation and identified potential key genes in the regulation of this process. These results will help to dissect the molecular mechanism of wood formation and provide key candidate genes for marker-assisted selection in B. luminifera.

Molecular Characterization of SQUAMOSA PROMOTER BINDING PROTEIN-LIKE (SPL) Gene Family in Betula luminifera.

As a major family of plant-specific transcription factors, SQUAMOSA PROMOTER BINDING PROTEIN-LIKE (SPL) genes play vital regulatory roles in plant growth, development and stress responses. In this study, 18 SPL genes were identified and cloned from Betula luminifera. Two zinc finger-like structures and a nuclear location signal (NLS) segments were existed in the SBP domains of all BlSPLs. Phylogenetic analysis showed that these genes were clustered into nine groups (group I-IX). The intron/exon structure and motif composition were highly conserved within the same group. 12 of the 18 BlSPLs were experimentally verified as the targets of miR156, and two cleavage sites were detected in these miR156-targeted BlSPL genes. Many putative cis-elements, associated with light, stresses and phytohormones response, were identified in the promoter regions of BlSPLs, suggesting that BlSPL genes are probably involved in important physiological processes and developmental events. Tissue-specific expression analysis showed that miR156-targeted BlSPLs exhibited a more differential expression pattern, while most miR156-nontargeted BlSPLs tended to be constitutively expressed, suggesting the distinct roles of miR156-targeted and nontargeted BlSPLs in development and growth of B. luminifera. Further expression analysis revealed that miR156-targeted BlSPLs were dramatically up-regulated with age, whereas mature BlmiR156 level was apparently declined with age, indicating that miR156/SPL module plays important roles in vegetative phase change of B. luminifera. Moreover, yeast two-hybrid assay indicated that several miR156-targeted and nontargeted BlSPLs could interact with two DELLA proteins (BlRGA and BlRGL), which suggests that certain BlSPLs take part in the GA regulated processes through protein interaction with DELLA proteins. All these results provide an important basis for further exploring the biological functions of BlSPLs in B. luminifera.