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The Physalis genus has long been used as traditional medicine in the treatment of various diseases. Physalins, the characteristic class of compounds in this genus, are major bioactive constituents. To date, the biogenesis of physalins remains largely unknown, except for the recently established knowledge that 24-methyldesmosterol is a precursor of physalin. To identify the genes encoding P450s that are putatively involved in converting 24-methyldesmosterol to physalins, a total of 306 P450-encoding unigenes were retrieved from our recently constructed P. angulata transcriptome. Extensive phylogenetic analysis proposed 21 P450s that might participate in physalin biosynthesis. To validate the candidates, we developed a virus-induced gene silencing (VIGS) system for P. angulata , and four P450 candidates were selected for the VIGS experiments. The reduction in the transcripts of the four P450 candidates by VIGS all led to decreased levels of physalin-class compounds in the P. angulata leaves. Thus, this study provides a number of P450 candidates that are likely associated with the biosynthesis of physalin-class compounds, forming a strong basis to reveal the unknown physalin biosynthetic pathway in the future.

The mechanisms underlying plant response to drought involve the expression of numerous functional and regulatory genes. Transcriptome sequencing based on the second- and/or third-generation high-throughput sequencing platforms has proven to be powerful for investigating the transcriptional landscape under drought stress. However, the full-length transcriptomes related to drought responses in the important conifer genus Pinus L. remained to be delineated using the third-generation sequencing technology. With the objectives of identifying the candidate genes responsible for drought and/or rehydration and clarifying the expression profile of key genes involved in drought regulation, we combined the third- and second-generation sequencing techniques to perform transcriptome analysis on seedling roots under drought stress and rewatering in the drought-tolerant conifer Pinus massoniana Lamb. A sum of 294,114 unique full-length transcripts were produced with a mean length of 3217 bp and N50 estimate of 5075 bp, including 279,560 and 124,438 unique full-length transcripts being functionally annotated and Gene Ontology enriched, respectively. A total of 4076, 6295 and 18,093 differentially expressed genes (DEGs) were identified in three pair-wise comparisons of drought-treatment versus control transcriptomes, including 2703, 3576 and 8273 upregulated and 1373, 2719 and 9820 downregulated DEGs, respectively. Moreover, 157, 196 and 691 DEGs were identified as transcription factors in the three transcriptome comparisons and grouped into 26, 34 and 44 transcription factor families, respectively. Gene Ontology enrichment analysis revealed that a remarkable number of DEGs were enriched in soluble sugar-related and cell wall-related processes. A subset of 75, 68 and 97 DEGs were annotated to be associated with starch, sucrose and raffinose metabolism, respectively, while 32 and 70 DEGs were associated with suberin and lignin biosynthesis, respectively. Weighted gene co-expression network analysis revealed modules and hub genes closely related to drought and rehydration. This study provides novel insights into root transcriptomic changes in response to drought dynamics in Masson pine and serves as a fundamental work for further molecular investigation on drought tolerance in conifers., (© The Author(s) 2023. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Aphidius gifuensis Ashmead is a dominant endoparasitoid of aphids, such as Myzus persicae and Sitobion avenae , and plays an important role in controlling aphids in various habitats, including tobacco plants and wheat in China. A. gifuensis has been successfully applied for the biological control of aphids, especially M. persicae , in green houses and fields in China. The corresponding parasites, as well as its mate-searching behaviors, are subjects of considerable interest. Previous A. gifuensis transcriptome studies have relied on short-read next-generation sequencing (NGS), and the vast majority of the resulting isotigs do not represent full-length cDNA. Here, we employed a combination of NGS and single-molecule real-time (SMRT) sequencing of virgin females (VFs), mated females (MFs), virgin males (VMs), and mated males (MMs) to comprehensively study the A. gifuensis transcriptome. Behavioral responses to the aphid alarm pheromone (E-β-farnesene, EBF) as well as to A. gifuensis of the opposite sex were also studied. VMs were found to be attracted by female wasps and MFs were repelled by male wasps, whereas MMs and VFs did not respond to the opposite sex. In addition, VFs, MFs, and MMs were attracted by EBF, while VMs did not respond. According to these results, we performed a personalized differential gene expression analysis of olfactory gene sets (66 odorant receptors, 25 inotropic receptors, 16 odorant-binding proteins, and 12 chemosensory proteins) in virgin and mated A. gifuensis of both sexes, and identified 13 candidate genes whose expression levels were highly consistent with behavioral test results, suggesting potential functions for these genes in pheromone perception. [ABSTRACT FROM AUTHOR]

Platysternon megacephalum is the sole living representative of the poorly studied turtle lineage Platysternidae. Their mitochondrial genome has been subject to gene rearrangement and control region duplication, resulting in a unique mitochondrial gene order in vertebrates. In this study, we sequenced the first full-length turtle (P. megacephalum) liver transcriptome using single-molecule real-time sequencing to study the transcriptional mechanisms of its mitochondrial genome. ND5 and ND6 anti-sense (ND6AS) forms a single transcript with the same expression in the human mitochondrial genome, but here we demonstrated differential expression of the rearranged ND5 and ND6AS genes in P. megacephalum. And some polycistronic transcripts were also reported in this study. Notably, we detected some novel long non-coding RNAs with alternative polyadenylation from the duplicated control region, and a novel ND6AS transcript composed of a long non-coding sequence, ND6AS, and tRNA-GluAS. These results provide the first description of a mtDNA transcriptome with gene rearrangement and control region duplication. These findings further our understanding of the fundamental concepts of mitochondrial gene transcription and RNA processing, and provide a new insight into the mechanism of transcription regulation of the mitochondrial genome. [ABSTRACT FROM AUTHOR]

Background: The flower of the safflower ( Carthamus tinctorius L.) has been widely used in traditional Chinese medicine for the ability to improve cerebral blood flow. Flavonoids are the primary bioactive components in safflower, and their biosynthesis has attracted widespread interest. Previous studies mostly used second-generation sequencing platforms to survey the putative flavonoid biosynthesis genes. For a better understanding of transcription data and the putative genes involved in flavonoid biosynthesis in safflower, we carry our study. Results: High-quality RNA was extracted from six types of safflower tissue. The RNAs of different tissues were mixed equally and used for multiple size-fractionated libraries (1–2, 2–3 and 3-6 k) library construction. Five cells were carried (2 cells for 1–2 and for 2-3 k libraries and 1 cell for 3-6 k libraries). 10.43Gb clean data and 38,302 de-redundant sequences were captured. 44 unique isoforms were annotated as encoding enzymes involved in flavonoid biosynthesis. The full length flavonoid genes were characterized and their evolutional relationship and expressional pattern were analyzed. They can be divided into eight families, with a large differences in the tissue expression. The temporal expressions under MeJA treatment were also measured, 9 genes are significantly up-regulated and 2 genes are significantly down-regulated. The genes involved in flavonoid synthesis in safflower were predicted in our study. Besides, the SSR and lncRNA are also analyzed in our study. Conclusions: Full-length transcriptome sequences were used in our study. The genes involved in flavonoid synthesis in safflower were predicted in our study. Combined the determination of flavonoids, CtC4H2 , CtCHS3 , CtCHI3 , CtF3H3 , CtF3H1 are mainly participated in MeJA promoting the synthesis of flavonoids. Our results also provide a valuable resource for further study on safflower. [ABSTRACT FROM AUTHOR]

In this study, we established a general framework to use PacBio full-length transcriptome sequencing for the investigation of mitochondrial RNAs. As a result, we produced the first full-length human mitochondrial transcriptome using public PacBio data and characterized the human mitochondrial genome with more comprehensive and accurate information. Other results included determination of the H-strand primary transcript, identification of the ND5/ND6AS/tRNA Glu AS transcript, discovery of palindrome small RNAs (psRNAs) and construction of the “mitochondrial cleavage” model, etc . These results reported for the first time in this study fundamentally changed annotations of human mitochondrial genome and enriched knowledge in the field of animal mitochondrial studies. The most important finding was two novel long non-coding RNAs (lncRNAs) of MDL1 and MDL1AS exist ubiquitously in animal mitochondrial genomes. [ABSTRACT FROM AUTHOR]

Selecting suitable plants as green manures is essential for improving marginal soil fertility and sustainable agroecosystems. Sophora alopecuroides is a highly salt- and alkali-tolerant leguminous plant that is often used as green manure. However, the molecular mechanism underlying saline-alkali tolerance in S. alopecuroides remains largely unclear. In this study, we investigated the molecular mechanisms underlying saline-alkali stress tolerance through physiological and transcriptomic analyses in S. alopecuroides seedlings. Comparative transcriptome analysis revealed that saline-alkali stress induced the expression of genes involved in cell wall modification, thereby increasing the contents of pectin and hemicellulose in roots and improving saline-alkali stress tolerance. The expression of genes related to the glycolysis pathway was induced, whereas genes involved in the TCA cycle and primary nitrogen metabolism were downregulated under salt and alkali stresses, indicating that S. alopecuroides plants modulate carbon flow to provide energy and ultimately regulate saline-alkali stress adaptation. Saline-alkali stress also modulated phytohormone signaling pathways in S. alopecuroides. Moreover, the leaves of seedlings under salt and alkali stresses contained elevated potassium, zinc and manganese contents, and the roots of seedlings under salt stress contained higher iron content. Taken together, these adaptive mechanisms to saline-alkali stress provide a physiological basis for the coordinated nutrient cycling of S. alopecuroides green manure. These results provide a molecular basis for the phytoremediation and utilization of saline-alkali soil, enabling the design of an integrated nutrient cycling system and optimization of the cultivation and field management of S. alopecuroides green manure. [Display omitted] • Saline-alkali stress modulates carbon flow to provide energy in S. alopecuroides. • Saline-alkali stress increases the contents of pectin and hemicellulose in roots. • Saline-alkali stress modulates phytohormone pathways. • Micronutrient contents were increased in S. alopecuroides under saline-alkali stress. • Improving saline-alkali land use efficiency by S. alopecuroides green manure. [ABSTRACT FROM AUTHOR]

[Display omitted] • A full-length transcriptional landscape of the silk gland in Antheraea pernyi was completed. • 7,568 genic loci were covered by 12,572 FL isoforms from the A. pernyi silk gland. • 1,121 lncRNAs and 871 TFs were first identified in the A. pernyi silk gland. • Some TF families underwent expansion or contraction of their members. Silk fibers spun by the silk gland of Antheraea pernyi have many unique properties and are of great value in genetic improvement and non-traditional applications. However, the complete transcriptional landscape and accurate genic annotation of the silk gland are yet to be conducted, which limits related studies on this organ. In this study, isoform sequencing revealed the full-length transcriptome of the A. pernyi silk gland, producing 12,572 high-confidence isoforms from 7,658 genes, among which more than 40 % of genes have not yet been annotated in the reference genome. Moreover, approximately 9 % of isoforms are computationally identified as long non-coding RNAs (lncRNAs). Up to 1,492 alternative splicing (AS) and 3,068 alternative polyadenylation (APA) events were revealed within this transcriptome. In addition, 2,569 putative transcription factors (TFs) belonging to 68 different families were first identified in A. pernyi genome, including 871 TFs in silk gland, and some TF families have undergone expansion or contraction. This study significantly improve our knowledge of the genes expressed in the silk gland of A. pernyi and provide a valuable resource for the in-depth study of silk protein synthesis and spinning, genetic improvement, and non-traditional applications in A. pernyi. [ABSTRACT FROM AUTHOR]

The genus Gnetum includes pantropical trees, shrubs and lianas, with unresolved phylogenetic relationships with other seed plant groups. Despite the reference genome for this genus being recently published, the molecular mechanisms that regulate the reproductive organ development of Gnetum remain unclear. A previous study showed that indole-3-acetic acid is involved in the regulation of female strobili of Gnetum, while the diversity and evolution of indole-3-acetic acid-related genes—the Aux/IAA genes—have never been investigated in Gnetales. Thus, a pooled sample from different developmental stages of female strobili in Gnetum luofuense C.Y. Cheng was sequenced using PacBio single-molecular long-read technology (SMRT) sequencing. PacBio SMRT sequencing generated a total of 53,057 full-length transcripts, including 2043 novel genes. Besides this, 10,454 alternative splicing (AS) events were detected with intron retention constituting the largest proportion (46%). Moreover, 1196 lncRNAs were identified, and 8128 genes were found to possess at least one poly (A) site. A total of 3179 regulatory proteins, including 1413 transcription factors (e.g., MADS-box and bHLHs), 477 transcription regulators (e.g., SNF2), and 1289 protein kinases (e.g., RLK/Pelles) were detected, and these protein regulators probably participated in the female strobili development of G. luofuense. In addition, this is the first study of the Aux/IAA genes of the Gnetales, and we identified 6, 7 and 12 Aux/IAA genes from Gnetum luofuense, Welwitschia mirabilis, and Ephedra equistina, respectively. Our phylogenetic analysis reveals that Aux/IAA genes from the gymnosperms tended to cluster and possessed gene structures as diverse as those in angiosperms. Moreover, the Aux/IAA genes of the Gnetales might possess higher molecular evolutionary rates than those in other gymnosperms. The sequencing of the full-length transcriptome paves the way to uncovering molecular mechanisms that regulate reproductive organ development in gymnosperms.

The planting of Pinus elliottii Engelm. has now reached close to three million ha in China. Molecular breeding as part of the improvement program for P. elliottii in southern China has been carried out in recent years. Third-generation sequencing (Pacbio sequencing technology, TGS) was used to obtain the exome of P. elliottii for molecular breeding. A total of 35.8 Gb clean reads were generated using TGS. After removing the redundant reads, we obtained 80,339 high-accuracy transcripts. Significantly, a total of 76,411 transcripts (95.1%) were blasted to public annotation databases. We predicted 65,062 intact coding sequences (CDSs), 8916 alternative splicing events, 1937 long non-coding RNAs, and 22,109 simple sequence repeats (SSRs) based on these obtained transcripts. Using the public databases and the data obtained above, 23 orthologous single-copy genes were identified to analyze the phylogenetic relationships for Pinus firstly including P. elliottii. Many positive selection genes involved in important biological processes and metabolism pathways were identified between P. elliottii and other pines. These positive selection genes could be candidate genes to be researched on the genetic basis of superior performance. Our study is the first to reveal the full-length and well-annotated transcripts of P. elliottii, which could provide reference for short transcriptome sequences in the research of genetics, phylogenetics, and genetic improvement for the non-reference genome species.

Sesuvium portulacastrum is a typical halophyte. However, few studies have investigated its salt-tolerant molecular mechanism. In this study, metabolome, transcriptome, and multi-flux full-length sequencing analysis were conducted to investigate the significantly different metabolites (SDMs) and differentially expressed genes (DEGs) of S. portulacastrum samples under salinity. The complete-length transcriptome of S. portulacastrum was developed, which contained 39,659 non-redundant unigenes. RNA-seq results showed that 52 DEGs involved in lignin biosynthesis may be responsible for S. portulacastrum salt tolerance. Furthermore, 130 SDMs were identified, and the salt response could be attributed to the p-coumaryl alcohol-rich in lignin biosynthesis. The co-expression network that was constructed after comparing the different salt treatment processes showed that the p-Coumaryl alcohol was linked to 30 DEGs. Herein, 8 structures genes, i.e., Sp4CL , SpCAD , SpCCR , SpCOMT , SpF5H , SpCYP73A , SpCCoAOMT , and SpC3′H were identified as significant factors in regulating lignin biosynthesis. Further investigation revealed that 64 putative transcription factors (TFs) may interact with the promoters of the above-mentioned genes. Together, the data revealed a potential regulatory network comprising important genes, putative TFs, and metabolites involved in the lignin biosynthesis of S. portulacastrum roots under salt stress, which could serve as a rich useful genetic resource for breeding excellent salt-tolerant plants. • The full-length transcriptome of Sesuvium portulacastrum roots was firstly obtained in this study. • The metabolites and genes involved in lignin biosynthesis played key roles in salt tolerance of S. portulacastrum. • p-Coumaryl alcohol might be the key contributors in S. portulacastrum salt tolerance. [ABSTRACT FROM AUTHOR]

The aim of this study was to elucidate the key targets of acupuncture in the colon of ulcerative colitis (UC) mice model using full-length transcriptome sequencing. 2.5% dextran sodium sulfate (DSS)-induced colitis mice were treated with or without acupuncture. Intestinal pathology was observed, and full transcriptome sequencing and bioinformatic analysis were performed. The results demonstrated that acupuncture treatment reduced the UC symptoms, disease activity index score, and histological colitis score and increased body weight, colon length, and the number of intestinal goblet cells. In addition, acupuncture can also decrease the expression of necrotic biomarker phosphorylates mixed lineage kinase domain-like pseudo kinase (p-MLKL). Full-length transcriptome analysis indicated that acupuncture reversed the expression of 987 of the 1918 upregulated differentially expressed genes (DEGs), and 632 of the 1351 downregulated DEGs induced by DSS. DEGs regulated by acupuncture were mainly involved in inflammatory responses and intestinal barrier pathways. The protein-protein interaction network analysis revealed that matrix metalloproteinases (MMPs) are important genes regulated by acupuncture. Gene set enrichment analysis revealed that extracellular matrix (ECM)-receptor interaction was an important target of acupuncture. In addition, alternative splicing analysis suggested that acupuncture improved signaling pathways related to intestinal permeability, the biological processes of xenobiotics, sulfur compounds, and that monocarboxylic acids are closely associated with MMPs. Overall, our transcriptome analysis results indicate that acupuncture improves intestinal barrier function in UC through negative regulation of MMPs expression., Competing Interests: Declaration of Conflicting InterestsThe author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

Although mitochondrial gene rearrangement has been observed in many insect lineages, little is known about how it affects mitochondrial gene transcription. To address this question, we first constructed a quantitative transcription map for Aphidius gifuensis , a species of parasitoid wasp known to have a highly rearranged mitochondrial genome (mitogenome) and two potential control regions (CRs). Based on this transcription map, we assessed the models of the mitochondrial transcription and post-transcription cleavage. We found that the J and N strand of this mitogenome differ significantly in transcriptional regulation. On the J strand, we found two transcription initiation sites (TISs), five transcription termination sites (TTSs), and six polycistronic primary transcripts whereas only one TIS, one TTS and one polycistronic primary transcript can be found on the N strand. Most of the non-coding regions of both strands were transcribed into primary transcripts and cleaved after transcription. The proposed mode of transcription of A. gifuensis was similar to that of Drosophila , a model organism with no gene rearrangement. And two rearranged gene clusters ( trnI-CR1-trnM-CR2-trnQ and trnW-trnY-trnC ) seemed to have little effects on the mode of transcription. In addition, our results revealed the presence of TISs in CR1 and CR2, implying that both CRs maybe required for transcriptional regulation. Analysis of the post-transcriptional cleavage process showed that there were both "forward cleavage" and "reverse cleavage" models in A. gifuensis , and more than one way of cleavages were found in three regions. The incomplete transcripts suggested that the direction of mitochondrial RNA degradation was from 5' to 3' end and supported the view of polyadenylation-dependent RNA degradation. Our study provides insights into the transcriptional and post-transcriptional regulation processes of highly rearranged insect mitogenomes., Competing Interests: The authors declare no conflict of interest., (© 2023 The Authors.)

• We performed full-length transcriptome sequencing of C. nitidissima flower. • 112 differentially expressed genes (DEGs), related to flavonoid synthesis, were identified by differential genes expression analysis. • Three R2R3-MYB and one bHLH that regulated flavonoid biosynthesis were screened out. • Transcriptional regulatory network of flavonoid biosynthesis in C. nitidissima Petals was constructed by WGCNA. Flavonoids are the main medicinal ingredients in Camellia nitidissima , but the regulatory mechanism of flavonoid biosynthesis in flowers is unclear; therefore, the flavonoids in C. nitidissima have not been effectively used. The present study performed full-length transcriptome sequencing of C. nitidissima flower. Furthermore, the reported RNA-sequencing data of C. nitidissima petals were reanalyzed using the full-length transcriptome as a reference, and the regulatory mechanism of flavonoid synthesis in petals was elucidated. The analysis identified 43,350 isoforms annotated in non-redundant protein (Nr), Kyoto Encyclopedia of Genes and Genomes (KEGG), EuKaryotic Orthologous Groups (KOG), and Swiss-Prot databases, among which 34,602 aligned to Camellia sinensis genes. A total of 11,857 differentially expressed genes (DEGs), including 112 related to flavonoid synthesis, were identified by pairwise comparison. Subsequently, analysis of the phylogeny and the conserved motifs of R2R3-MYB using the proteins sequences identified three R2R3-MYB transcription factors that regulated flavonoid biosynthesis. Weighted gene co-expression network analysis (WGCNA) identified phenylalanine ammonia-lyase (PAL) and 4-coumarate: CoA ligase (4CL) as the hub genes and showed that bHLH79 interacted with PAL. Finally, validated the expression of seven DEGs involved in flavonoid biosynthesis using real-time quantitative PCR (qRT-PCR). Thus, the present study generated and used the full-length transcriptome as the reference to analyze the transcriptome of petals and proposed a possible regulatory mechanism of flavonoid synthesis in C. nitidissima. The study's findings unravel the genetic mechanisms underlying flavonoid synthesis and suggest candidate genes for the genetic improvement of C. nitidissima. [ABSTRACT FROM AUTHOR]

As the carrier of genetic information, RNA carries the information from genes to proteins. Transcriptome sequencing technology is an important way to obtain transcriptome sequences, and it is also the basis for transcriptome research. With the development of third-generation sequencing, long reads can cover full-length transcripts and reflect the composition of different isoforms. However, the high error rate of third-generation sequencing affects the accuracy of long reads and downstream analysis. The current error correction methods seldom consider the existence of different isoforms in RNA, which makes the diversity of isoforms a serious loss. Here, we introduce LCAT (long-read error correction algorithm for transcriptome sequencing data), a wrapper algorithm of MECAT, to reduce the loss of isoform diversity while keeping MECAT's error correction performance. The experimental results show that LCAT can not only improve the quality of transcriptome sequencing long reads but also retain the diversity of isoforms., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Zhu and Liao.)

Cold stress impairs plant growth and development, resulting in crop failure. Cultivated potato ( Solanum tuberosum L.) is sensitive to freezing, while its wild relative, S. commersonii , has a strong freezing tolerance. To decipher the anti-freezing mechanism of CM, we carried out a transcriptomic and metabolomic analysis of an anti-freezing variety of CM (a type of S. commersonii ) and a freeze-sensitive variety of DM (a type of Solanum tuberosum L.). A total of 49,232 high-quality transcripts from 12,811 gene loci, including 46,772 coding sequences and 2018 non-coding RNAs, were identified. KEEG enrichment analysis of differentially expressed genes (DEGs) between the two varieties showed that the flavonoid biosynthesis pathway was strongly induced by freezing stress, which was proven by flavonoid metabolome analysis. Consistent with the accumulation of more flavonoids, nearly all the pathway genes were significantly upregulated in CM than those in DM. The transcript levels of two chalcone synthase ( CHS - 1 ) isoforms and four isoforms of flavonoid 3'-hydroxylase ( F3'H - 1 ) were confirmed by qRT-PCR. Co-expression analysis identified one Myb-related and three UGT s ( UDP - glycosyltransferase ) that were significantly upregulated in CM during freezing stress. Our findings support that the flavonoid pathway was significantly enhanced by freezing stress and the greater accumulation ofglycosylatedflavonoids in resistant types than that of sensitive types, maybe accounting for the increased freezing tolerance of freeze-resistant potato varieties.

Kentucky bluegrass ( Poa pratensis L.) is an eminent turfgrass species with a complex genome, but it is sensitive to rust ( Puccinia striiformis ). The molecular mechanisms of Kentucky bluegrass in response to rust still remain unclear. This study aimed to elucidate differentially expressed lncRNAs (DELs) and genes (DEGs) for rust resistance based on the full-length transcriptome. First, we used single-molecule real-time sequencing technology to generate the full-length transcriptome of Kentucky bluegrass. A total of 33,541 unigenes with an average read length of 2,233 bp were obtained, which contained 220 lncRNAs and 1,604 transcription factors. Then, the comparative transcriptome between the mock-inoculated leaves and rust-infected leaves was analyzed using the full-length transcriptome as a reference genome. A total of 105 DELs were identified in response to rust infection. A total of 15,711 DEGs were detected (8,278 upregulated genes, 7,433 downregulated genes) and were enriched in plant hormone signal transduction and plant-pathogen interaction pathways. Additionally, through co-location and expression analysis, it was found that lncRNA56517, lncRNA53468, and lncRNA40596 were highly expressed in infected plants and upregulated the expression of target genes AUX/IAA , RPM1 , and RPS2 , respectively; meanwhile, lncRNA25980 decreased the expression level of target gene EIN3 after infection. The results suggest that these DEGs and DELs are important candidates for potentially breeding the rust-resistant Kentucky bluegrass., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Zhao, Sun, Chen, Wu, Liu, Jiang, Xie and Chen.)

Zanthoxylum nitidum (Roxb.) DC. ( Z. nitidum ) is a type of Chinese Dao-di herb, also called Liangmianzhen, which is widely used to treat arthralgia, rheumatic arthralgia, and stomach pain. However, genomic resources for Z. nitidum are still scarce. This study provides transcriptomic resources for Z. nitidum by applying single-molecule real-time (SMRT) sequencing technology. In total, 456,109 circular consensus sequencing (CCS) reads were generated with a mean length of 2,216 bp from Z. nitidum roots, old stems, young branches, leaves, flowers, and fruits. Of these total reads, 353,932 were full-length nonchimeric (FLNC) reads with an average length of 1,996 bp. A total of 16,163 transcripts with a mean length of 1,171 bp were acquired. Of these transcripts, 14,231 (88%) were successfully annotated using public databases. Across all the 16,163 transcripts, we identified 6,255 long non-coding RNAs (lncRNAs) and 22,780 simple sequence repeats (SSRs). Furthermore, 3,482 transcription factors were identified. Among the SSR loci, 1-3 nucleotide repeats were dominant, occupying 99.36% of the total SSR loci, with mono-, di-, and tri-nucleotide repeats accounting for 61.80%, 19.89%, and 5.02% of the total SSR loci, respectively. A total of 36 out of 100 randomly selected primer pairs were verified to be positive, 20 of which showed polymorphism. These findings enrich the genetic resources available for facilitating future studies and research on relevant topics such as population genetics in Z. nitidum ., Competing Interests: Zhu Yanxia, Huang Yanfen, Wei Kunhua and Jiang Jianping are all scientific researchers of Guangxi Medical Botanical Garden. Junnan Yu is employed by ChongQing Jinzhi Quality Certification Co., LTD. The authors declare that they have no competing interests., (© 2023 Zhu et al.)

Background: The extremely harsh environment of the desert is changing dramatically every moment, and the rapid adaptive stress response in the short term requires enormous energy expenditure to mobilize widespread regulatory networks, which is all the more detrimental to the survival of the desert plants themselves. The dune reed, which has adapted to desert environments with complex and variable ecological factors, is an ideal type of plant for studying the molecular mechanisms by which Gramineae plants respond to combinatorial stress of the desert in their natural state. But so far, the data on the genetic resources of reeds is still scarce, therefore most of their research has focused on ecological and physiological studies., Results: In this study, we obtained the first De novo non-redundant Full-Length Non-Chimeric (FLNC) transcriptome databases for swamp reeds (SR), dune reeds (DR) and the All of Phragmites australis (merged of iso-seq data from SR and DR), using PacBio Iso-Seq technology and combining tools such as Iso-Seq3 and Cogent. We then identified and described long non-coding RNAs (LncRNA), transcription factor (TF) and alternative splicing (AS) events in reeds based on a transcriptome database. Meanwhile, we have identified and developed for the first time a large number of candidates expressed sequence tag-SSR (EST-SSRs) markers in reeds based on UniTransModels. In addition, through differential gene expression analysis of wild-type and homogenous cultures, we found a large number of transcription factors that may be associated with desert stress tolerance in the dune reed, and revealed that members of the Lhc family have an important role in the long-term adaptation of dune reeds to desert environments., Conclusions: Our results provide a positive and usable genetic resource for Phragmites australis with a widespread adaptability and resistance, and provide a genetic database for subsequent reeds genome annotation and functional genomic studies., (© 2023. The Author(s).)

Potato virus Y (PVY) mainly infects Solanaceous crops, resulting in considerable losses in the yield and quality. Iron (Fe) is involved in various biological processes in plants, but its roles in resistance to PVY infection has not been reported. In this study, foliar application of Fe could effectively inhibit early infection of PVY, and a full-length transcriptome and Illumina RNA sequencing was performed to investigate its modes of action in PVY-infected Nicotiana tabacum . The results showed that 18,074 alternative splicing variants, 3,654 fusion transcripts, 3,086 long non-coding RNAs and 14,403 differentially expressed genes (DEGs) were identified. Specifically, Fe application down-regulated the expression levels of the DEGs related to phospholipid hydrolysis, phospholipid signal, cell wall biosynthesis, transcription factors (TFs) and photosystem I composition, while those involved with photosynthetic electron transport chain (PETC) were up-regulated at 1 day post inoculation (dpi). At 3 dpi, these DEGs related to photosystem II composition, PETC, molecular chaperones, protein degradation and some TFs were up-regulated, while those associated with light-harvesting, phospholipid hydrolysis, cell wall biosynthesis were down-regulated. At 9 dpi, Fe application had little effects on resistance to PVY infection and transcript profiles. Functional analysis of these potentially critical DEGs was thereafter performed using virus-induced gene silencing approaches and the results showed that NbCat-6A positively regulates PVY infection, while the reduced expressions of NbWRKY26 , NbnsLTP , NbFAD3 and NbHSP90 significantly promote PVY infection in N. benthamiana . Our results elucidated the regulatory network of Fe-mediated resistance to PVY infection in plants, and the functional candidate genes also provide important theoretical bases to further improve host resistance against PVY infection., Competing Interests: Authors CX, YZ and QX were employed by company Luzhou City Company of Sichuan Province Tobacco Company. Authors DZ and QM were employed by company Guizhou Qianxinan Prefectural Tobacco Company. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Xu, Guo, Li, Lan, Zhang, Xie, Zhu, Mu, Wang, An, Xia and Wu.)

Viruses deploy numerous strategies to infect plants, typically by forming complexes with another virus, leading to more efficient infection. However, the detailed plant responses to viral infection and the underlying mechanisms of co-infection remain unclear. Previously, we found that tomato spotted wilt orthotospovirus (TSWV) and Hippeastrum chlorotic ringspot orthotospovirus (HCRV) could infect plants in the field by forming a complex. In this study, we found that TSWV infected tobacco ( Nicotiana benthamiana ) plants in cooperation with HCRV, leading to a more efficient infection rate of both viruses. We then used the in-depth full-length transcriptome to analyze the responses of N. benthamiana to complex infection by TSWV-HCRV (TH). We found that infection with individual TSWV and HCRV triggered plant defense responses, including the jasmonic acid signaling pathway, autophagy, and secondary metabolism. However, TH co-infection could not trigger and even suppress some genes that are involved in these basal resistance responses, suggesting that co-infection is advantageous for the virus and not for the plants. Typically, the TH complex inhibits NbPR1 expression to suppress tobacco resistance. Moreover, the TH complex could alter the expression of microRNAs (miRNAs), especially novel-m0782-3p and miR1992-3p, which directly interact with NbSAM and NbWRKY6 and suppress their expression in tobacco, leading to downregulation of NbPR1 and loss of resistance in tobacco to TSWV and HCRV viruses. Overall, our results elucidated the co-infection mechanisms of TH in tobacco by deploying the miRNA of plants to suppress plant basal resistance and contributed to developing a novel strategy to control crop disease caused by this virus complex., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Gui, Hu, Jia, Gao, Tao, Li and Liu.)

Objectives: Diospyros oleifera, one of the most economically important Diospyros species, is an ideal model for studying the fruit development of persimmon. While, the lack of whole-transcriptome has hindered the complex transcriptional regulation mechanisms of sugar and tannin during fruit development., Data Description: We applied Oxford Nanopore Technologies to six developmental stage of fruit from D. oleifera for use in transcriptome sequencing. As a result of full-length transcriptome sequencing, 55.87 Gb of clean data were generated. After mapping onto the reference genome of D. oleifera, 51,588 full-length collapsing transcripts, including 2,727 new gene loci and 43,223 transcripts, were obtained. Comprehensively annotated, 38,086 of new transcripts were functional annotation, and 972 lncRNAs, 7,159 AS events were predicted. Here, we released the transcriptome database of D. oleifera at different stage of fruit development,which will provide a fundamention of to investigatethe transcript structure, variants and evolution of persimmon., (© 2023. The Author(s).)

The Great Himalayan Leaf-nosed bat ( Hipposideros armiger ) is one of the most representative species of all echolocating bats and is an ideal model for studying the echolocation system of bats. An incomplete reference genome and limited availability of full-length cDNAs have hindered the identification of alternatively spliced transcripts, which slowed down related basic studies on bats' echolocation and evolution. In this study, we analyzed five organs from H. armiger for the first time using PacBio single-molecule real-time sequencing (SMRT). There were 120 GB of subreads generated, including 1,472,058 full-length non-chimeric (FLNC) sequences. A total of 34,611 alternative splicing (AS) events and 66,010 Alternative Polyadenylation (APA) sites were detected by transcriptome structural analysis. Moreover, a total of 110,611 isoforms were identified, consisting of 52% new isoforms of known genes and 5% of novel gene loci, as well as 2112 novel genes that have not been annotated before in the current reference genome of H. armiger . Furthermore, several key novel genes, including Pol , RAS , NFKB1 , and CAMK4 , were identified as being associated with nervous, signal transduction, and immune system processes, which may be involved in regulating the auditory nervous perception and immune system that helps bats to regulate in echolocation. In conclusion, the full-length transcriptome results optimized and replenished existing H. armiger genome annotation in multiple ways and offer advantages for newly discovered or previously unrecognized protein-coding genes and isoforms, which can be used as a reference resource.

• In the present study, the assay for biochemical and molecular mechanism were used by 13C labeling, full-length RNA-seq and gene transformation. • We make a conclusion that 50%FL significantly improved the source and sink strength. As a prime candidate gene, overexpression of VvPUP1 in grape cells improved its sugar accumulation. • These results demonstrates that the optimal fruit load was about 50%. Appropriately adjusting fruit loads (FL) can promote plant productivity in the field. In the present study, physiological, transcriptomic, and gene function assays were conducted by 13C labeling, full-length RNA-seq, and gene transformation. A comparative analysis of three different FL treatments (25%FL, 50%FL, and 100% FL) was performed to identify the mechanism that response to different FL. The results showed that 50%FL significantly improved the source strength by enhancing photosynthesis. The sink strength of 50%FL also reached the maximum when considering soluble sugar, weight, 13C allocation, sugar metabolism-related enzyme activity, and composition in berries, and yield. In addition, full-length transcriptome profiling exploited 1,098 differently expressed genes (DEGs) in 25%FL and 100%FL, and 1,479 DEGs in 50%FL and 100%FL, respectively. Thirty-four genes, including PUP1 and FBA1 , were identified in carbon transport and metabolism-related pathways. As a major candidate gene, VvPUP1 was found to enhance sugar accumulation when overexpressed in grape cells. The transgenic cells had high soluble sugar contents and sucrose synthase (SS), neutral invertase (NI), and sucrose phosphate synthase (SPS) activities. Overexpression of VvPUP1 in tomatoes increased fruit size and weight, soluble sugar concentration, and the expression levels of SlSUS1, SlSPS1 , and SlBGLU13 , suggesting that VvPUP1 can promote berry enlargement and sugar accumulation. These results provide novel insights into the response of grapevine to different FL, demonstrating the differences in the source (leaf) and sink (berry) strength among different FL grapes. [ABSTRACT FROM AUTHOR]

A new genome editing system named NgAgo-gDNA was invented using 5，phosphorylated single-stranded guide DNA (gDNA) of 24 nucleotides and Natronobacterium gregoryi Argonaute (NgAgo). This system outperformed the RNA-guided genome editing system CRISPR-Cas9 on several features. The success of the NgAgogDNA project demonstrated the importance of bioinformatics in biological research and will stimulate the development of genome editing systems. The NgAgo-gDNA project was initiated from searching homologs of TtAgo and PfAgo，two other enzymes from the AGO protein family. The authors used the software PSI-BLAST against the NCBI NR database to retrieve homologous protein sequences. After further analysis and filtering， they found the NgAgo protein (GenBank ： AFZ73749.1)，which works at the temperature of 37 冗.The key step in the NgAgogDNA project is to narrow down a great number of AGO homologous protein sequences to several candidates using bioinformatics methods for experimental validation of their functions. These bioinformatics methods were not explained in the published paper but could belong to the empirical methodology. An alternative but advanced methodology is to use machine learning algorithms (e.g. support vector machine or random forest) to modify AGO proteins which work at a temperature close to 37 冗.The future studies can be conducted in several fields using bioinformatics methods. First，the structural information of the NgAgo protein can be used to reveal the mechanism of the DNA and protein interaction. The sequence with structure comparison between NgAgo and TtAgo & PfAgo or other AGO proteins will help understand their molecular functions. Second， using the sequence or structure similarities，more RNA-or DNA-binding proteins can be retrieved from the public databases to help design new genome editing systems. Third，since RNAi (RNA interference) uses AGO to cleave double stranded RNAs，the guide-target complexes of AGO proteins need be studied to reveal the common mechanisms and differences between genome editing and RNAi. Fourth，a great number ol AGO genes from lower to higher organisms can be used to study the evolution ol AGO and the coevolution between the viruses and the hosts. [ABSTRACT FROM AUTHOR]

Hyphantria cunea (Drury) is a globally important forest pest. We found that the Serratia marcescens Bizio strain SM1 had insecticidal activity against H. cunea , but the transcriptomic response of H. cunea to SM1 were not clear. Therefore, we performed full-length sequencing of the transcriptomes of H. cunea larvae infected with SM1 and the control group. A total of 1,183 differentially expressed genes (DEGs) were identified by comparing the group infected with SM1 and the control group, including 554 downregulated genes and 629 upregulated genes. We found many downregulated genes in metabolic pathways. Furthermore, some of these downregulated genes were involved in cellular immunity, melanization, and detoxification enzymes, which showed that SM1 weakened H. cunea immunity. In addition, genes in the juvenile hormone synthesis pathway were upregulated, which was detrimental to the survival of H. cunea . This research analyzed the transcriptomic response of H. cunea to SM1 by high-throughput full-length transcriptome sequencing. The results provide useful information to explore the relationship between S. marcescens and H. cunea , and theoretical support for the application of S. marcescens and the control of H. cunea in the future., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Zhang, Tang, Wang and Tang.)

Selenium (Se) is an essential element for mammals, and its deficiency in the diet is a global problem. Agronomic biofortification through exogenous Se provides a valuable strategy to enhance human Se intake. Selenium nanoparticles (SeNPs) have been regarded to be higher bioavailability and less toxicity in comparison with selenite and selenate. Still, little has been known about the mechanism of their metabolism in plants. Soybean ( Glycine max L.) can enrich Se, providing an ideal carrier for Se biofortification. In this study, soybean sprouts were treated with SeNPs, and a combination of next-generation sequencing (NGS) and single-molecule real-time (SMRT) sequencing was applied to clarify the underlying molecular mechanism of SeNPs metabolism. A total of 74,662 nonredundant transcripts were obtained, and 2109 transcription factors, 9687 alternative splice events, and 3309 long non-coding RNAs (lncRNAs) were predicted, respectively. KEGG enrichment analysis of the DEGs revealed that metabolic pathways, biosynthesis of secondary metabolites, and peroxisome were most enriched both in roots and leaves after exposure to SeNPs. A total of 117 transcripts were identified to be putatively involved in SeNPs transport and biotransformation in soybean. The top six hub genes and their closely coexpressed Se metabolism-related genes, such as adenylylsulfate reductase ( APR3 ), methionine-tRNA ligase ( SYM ), and chloroplastic Nifs-like cysteine desulfurases ( CNIF1 ), were screened by WGCNA and identified to play crucial roles in SeNPs accumulation and tolerance in soybean. Finally, a putative metabolism pathway of SeNPs in soybean was proposed. These findings have provided a theoretical foundation for future elucidation of the mechanism of SeNPs metabolism in plants.