De Novo Transcriptome Analysis Research Articles

De Novo Transcriptome Analysis by PacBio SMRT-Seq and Illumina RNA-Seq Provides New Insights into Polyphenol Biosynthesis in Chinese Olive Fruit

Polyphenols play a crucial role in fruit flavor. To elucidate the mechanism of fruit polyphenol metabolism, we constructed a transcriptome atlas through PacBio single-molecule real-time (SMRT) sequencing and Illumina next-generation sequencing (NGS) using Canarium album (Lour.) Raeusch., which is a fantastic fruit rich in polyphenolic compounds. In this work, PacBio full-length transcriptome assembly generated 135,439 isoforms with an average length of all isoforms of 2687.94 bp and an N50 length of 3224 bp. To gain deeper insights into the molecular mechanisms of polyphenol biosynthesis in C. album, we constructed twelve RNA-Seq libraries from four developmental stages of the fruits. We identified a total of 28,658 differentially expressed genes (DEGs). We found that many DEGs were involved in metabolic pathways, biosynthesis of secondary metabolites, biosynthesis of antibiotics, starch and sucrose metabolism, and plant hormone signal transduction. Here, we report the expression profiles of 215 DEGs encoding 27 enzymes involved in the polyphenol biosynthesis pathway in C. album. In addition, 285 differentially expressed transcription factors (TFs) continuously down-regulated in four developmental periods of C. album fruit, which may indicate their potential role in the response to polyphenol metabolism and phenylpropanoid biosynthesis pathways. This report will help us understand polyphenol biosynthesis’s functions and metabolic mechanism in C. album. The transcriptome data provide a valuable resource for genetic and genomics research. They will facilitate future work exploiting C. album and other fruits used as medicine and food.

De Novo RNA Sequencing and Transcriptome Analysis of Sclerotium rolfsii Gene Expression during Sclerotium Development.

Sclerotium rolfsii is a destructive soil-borne fungal pathogen that causes stem rot in cultivated plants. However, little is known about the genetic basis of sclerotium development. In this study, we conducted de novo sequencing of genes from three different stages of S. rolfsii (mycelia, early sclerotium formation, and late sclerotium formation) using Illumina HiSeqTM 4000. We then determined differentially expressed genes (DEGs) across the three stages and annotated gene functions. STEM and weighted gene-co-expression network analysis were used to cluster DEGs with similar expression patterns. Our analysis yielded an average of 25,957,621 clean reads per sample (22,913,500-28,988,848). We identified 8929, 8453, and 3744 DEGs between sclerotium developmental stages 1 versus 2, 1 versus 3, and 2 versus 3, respectively. Additionally, four significantly altered gene expression profiles involved 220 genes related to sclerotium formation, and two modules were positively correlated with early and late sclerotium formation. These results were supported by the outcomes of qPCR and RNA-sequencing conducted on six genes. This is the first study to provide a gene expression map during sclerotial development in S. rolfsii, which can be used to reduce the re-infection ability of this pathogen and provide new insights into the scientific prevention and control of the disease. This study also provides a useful resource for further research on the genomics of S. rolfsii.

De-novo transcriptome assembly and analysis of lettuce plants grown under red, blue or white light

Lettuce (Lactuca sativa) is grown in various parts of the world for use as a leafy vegetable. Although the use of light-emitting diode (LED) in controlled plant production systems has been successfully used to enhance nutritional quality and plant growth efficiently, the molecular basis of lettuce’s response to varying light spectra is not studied. Using next-generation sequencing, we have analyzed the transcriptomes of leaf lettuce (Lactuca sativa var. ‘New Red Fire’) grown hydroponically in a modular agricultural production system under three different types of LED lighting: red, blue, and white light. Illumina HiSeq sequencing platform was used to generate paired-end sequence reads (58 Gb raw and 54 Gb clean data) of the transcriptome of lettuce leaves exposed to varying light spectra. The de novo assembled final transcriptome contained 74,096 transcripts. Around 53% and 39% of the assembled transcripts matched to the UniProt and RefSeq RNA sequences, respectively. The validation of the differentially expressed transcripts using RT-qPCR showed complete agreement with RNA-Seq data for 27 transcripts. A comparison of the blue versus red light treatments showed the highest number of significantly differentially expressed transcripts. Among the transcripts significantly up-regulated in blue-light-exposed leaves compared to white-light-exposed leaves, ~ 26% were involved in the ‘response to stress’. Among the transcripts significantly upregulated under red light compared to white light, ~ 6% were associated with ‘nucleosome assembly’ and other processes, such as ‘oxidation–reduction process’ and ‘response to water deprivation’ were significantly enriched. Thus, the result from the current study provides deeper insights into differential gene expression patterns and associated functional aspects under varying light qualities.

De-novo transcriptome analysis unveils differentially expressed genes regulating drought and salt stress response in Panicum sumatrense

Screening the transcriptome of drought tolerant variety of little millet (Panicum sumatrense), a marginally cultivated, nutritionally rich, susbsistent crop, can identify genes responsible for its hardiness and enable identification of new sources of genetic variation which can be used for crop improvement. RNA-Seq generated ~ 230 million reads from control and treated tissues, which were assembled into 86,614 unigenes. In silico differential gene expression analysis created an overview of patterns of gene expression during exposure to drought and salt stress. Separate gene expression profiles for leaf and root tissue revealed the differences in regulatory mechanisms operating in these tissues during exposure to abiotic stress. Several transcription factors were identified and studied for differential expression. 61 differentially expressed genes were found to be common to both tissues under drought and salinity stress and were further validated using qRT-PCR. Transcriptome of P. sumatrense was also used to mine for genic SSR markers relevant to abiotic stress tolerance. This study is first report on a detailed analysis of molecular mechanisms of drought and salinity stress tolerance in a little millet variety. Resources generated in this study can be used as potential candidates for further characterization and to improve abiotic stress tolerance in food crops.

In silico identification of effector proteins from generalist herbivore Spodoptera litura

BackgroundThe common cutworm, Spodoptera litura Fabricius is a leaf and fruit feeding generalist insect of the order Lepidoptera and a destructive agriculture pest. The broad host range of the herbivore is due to its ability to downregulate plant defense across different plants. The identity of Spodoptera litura released effectors that downregulate plant defense are largely unknown. The current study aims to identify genes encoding effector proteins from salivary glands of S. litura (Fab.).ResultsHead and salivary glands of Spodoptera litura were used for de-novo transcriptome analysis and effector prediction. Eight hundred ninety-nine proteins from the head and 330 from salivary gland were identified as secretory proteins. Eight hundred eight proteins from the head and 267 from salivary gland proteins were predicted to be potential effector proteins.ConclusionsThis study is the first report on identification of potential effectors from Spodoptera litura salivary glands.

Denovo transcriptome analysis and gene expression profiling of fish scales isolated from Carassiusauratus during space flight: Impact of melatonin on gene expression in response to space radiation.

Astronauts are inevitably exposed to two major risks during space flight, microgravity and radiation. Exposure to microgravity has been discovered to lead to rapid and vigorous bone loss due to elevated osteoclastic activity. In addition, long-term exposure to low-dose-rate space radiation was identified to promote DNA damage accumulation that triggered chronic inflammation, resulting in an increased risk for bone marrow suppression and carcinogenesis. In our previous study, melatonin, a hormone known to regulate the sleep-wake cycle, upregulated calcitonin expression levels and downregulated receptor activator of nuclear factor-κB ligand expression levels, leading to improved osteoclastic activity in a fish scale model. These results indicated that melatonin may represent a potential drug or lead compound for the prevention of bone loss under microgravity conditions. However, it is unclear whether melatonin affects the biological response induced by space radiation. The aim of the present study was to evaluate the effect of melatonin on the expression levels of genes responsive to space radiation. In the present study, to support the previous data regarding de novo transcriptome analysis of goldfish scales, a detailed and improved experimental method (e.g., PCR duplicate removal followed by de novo assembly, global normalization and calculation of statistical significance) was applied for the analysis. In addition, the transcriptome data were analyzed via global normalization, functional categorization and gene network construction to determine the impact of melatonin on gene expression levels in irradiated fish scales cultured in space. The results of the present study demonstrated that melatonin treatment counteracted microgravity- and radiation-induced alterations in the expression levels of genes associated with DNA replication, DNA repair, proliferation, cell death and survival. Thus, it was concluded that melatonin may promote cell survival and ensure normal cell proliferation in cells exposed to space radiation.

De Novo Transcriptome Analysis of Durum Wheat Flag Leaves Provides New Insights Into the Regulatory Response to Elevated CO2 and High Temperature.

Global warming is becoming a significant problem for food security, particularly in the Mediterranean basin. The use of molecular techniques to study gene-level responses to environmental changes in non-model organisms is increasing and may help to improve the mechanistic understanding of durum wheat response to elevated CO2 and high temperature. With this purpose, we performed transcriptome RNA sequencing (RNA-Seq) analyses combined with physiological and biochemical studies in the flag leaf of plants grown in field chambers at ear emergence. Enhanced photosynthesis by elevated CO2 was accompanied by an increase in biomass and starch and fructan content, and a decrease in N compounds, as chlorophyll, soluble proteins, and Rubisco content, in association with a decline of nitrate reductase and initial and total Rubisco activities. While high temperature led to a decline of chlorophyll, Rubisco activity, and protein content, the glucose content increased and starch decreased. Furthermore, elevated CO2 induced several genes involved in mitochondrial electron transport, a few genes for photosynthesis and fructan synthesis, and most of the genes involved in secondary metabolism and gibberellin and jasmonate metabolism, whereas those related to light harvesting, N assimilation, and other hormone pathways were repressed. High temperature repressed genes for C, energy, N, lipid, secondary, and hormone metabolisms. Under the combined increases in atmospheric CO2 and temperature, the transcript profile resembled that previously reported for high temperature, although elevated CO2 partly alleviated the downregulation of primary and secondary metabolism genes. The results suggest that there was a reprogramming of primary and secondary metabolism under the future climatic scenario, leading to coordinated regulation of C-N metabolism towards C-rich metabolites at elevated CO2 and a shift away from C-rich secondary metabolites at high temperature. Several candidate genes differentially expressed were identified, including protein kinases, receptor kinases, and transcription factors.

Differential Regulation of Anthocyanins in Green and Purple Turnips Revealed by Combined De Novo Transcriptome and Metabolome Analysis.

Purple turnip Brassica rapa ssp. rapa is highly appreciated by consumers but the metabolites and molecular mechanisms underlying the root skin pigmentation remain open to study. Herein, we analyzed the anthocyanin composition in purple turnip (PT) and green turnip (GT) at five developmental stages. A total of 21 anthocyanins were detected and classified into the six major anthocynanin aglycones. Distinctly, PT contains 20 times higher levels of anthocyanins than GT, which explain the difference in the root skin pigmentation. We further sequenced the transcriptomes and analyzed the differentially expressed genes between the two turnips. We found that PT essentially diverts dihydroflavonols to the biosynthesis of anthocyanins over flavonols biosynthesis by strongly down-regulating one flavonol synthase gene, while strikingly up-regulating dihydroflavonol 4-reductase (DFR), anthocyanidin synthase and UDP-glucose: flavonoid-3-O-glucosyltransferase genes as compared to GT. Moreover, a nonsense mutation identified in the coding sequence of the DFR gene may lead to a nonfunctional protein, adding another hurdle to the accumulation of anthocyanin in GT. We also uncovered several key members of MYB, bHLH and WRKY families as the putative main drivers of transcriptional changes between the two turnips. Overall, this study provides new tools for modifying anthocyanin content and improving turnip nutritional quality.

Comparative De Novo transcriptome analysis of the Australian black-lip and Sydney rock oysters reveals expansion of repetitive elements in Saccostrea genomes.

Ostreid oysters (the ‘true oysters’) represent a large and commercially important family of bivalve molluscs. Several species, such as the Pacific oyster (Magallana gigas), the American oyster (Crassostrea virginica), the European oyster (Ostrea edulis) and the Sydney rock oyster (Saccostrea glomerata), are currently farmed at a large scale. However a number of other species may also be suitable for commercial-scale aquaculture. One such species is the ‘black-lip oyster’, a large Saccostrea species of uncertain taxonomic affinity found in northern Australia. Here, phylogenetic analysis of the COI gene places this oyster within a clade identified in a previous study of Japanese Saccostrea species, ‘Saccostrea lineage J’. To facilitate comparisons between this oyster and the better-studied S. glomerata, de novo transcriptomes were generated from larval stages and adult tissues of both species. Patterns of orthology indicated an expansion of repetitive elements within Saccostrea genomes when compared to M. gigas and C. virginica, which may be reflected in increased evolutionary rates and/or genome sizes. The generation of high-quality transcriptomes for these two commercially relevant oysters provides a valuable resource for gene identification and comparison of molecular processes in these and other mollusc species.

De Novo Transcriptome Sequencing and Analysis to Identify Genes Involved in the Biosynthesis of Flavonoids in Abrus mollis Leaves

Abrus mollis Hance is an endemic genuine medicinal material growing widely in Southern China. We used the next-generation sequencing technology to carry out de novo transcriptome sequencing and analysis of A. mollis leaves. 57669118 high quality reads and 53743 assembled unigenes were obtained. 36271 unigenes (67.49%) were annotated in the four public databases (NCBI-nr, Swiss-Prot, KEGG, and KOG). After comprehensive analysis of the A. mollis transcriptome, 35656 unigenes were classified into 25 KOG function classifications and 7572 unigenes were assigned to 128 different KEGG pathways. Then, the unigenes with their key enzymes in phenylalanine, phenylpropanoid, flavonoids and isoflavonoid biosynthesis pathway were investigated in KEGG pathways. The study results will be helpful to reveal the biosynthesis mechanism of flavonoids and isoflavonoid in A. mollis leaves and carry out its biological regulation research.

De-novo assembly and transcriptome analysis of Odontesthes argentinensis gill tissue, with development of single sequence repeat markers

One of the most common transitions observed in actinopterygian fishes were from marine to freshwater environments. This phenomenon could suggest that osmoregulatory mechanisms in this group are particularly adaptable to salinity changes. The silversides of genus Odontesthes are not the exception within Atherinomorpha, with several invasions from marine to freshwater environments reported for this group. In this work, a de-novo transcriptome assembly of Odonthestes argentinensis gill tissue was performed, by means of paired-end sequencing in an Illumina HiSeq 2000 platform. Analyses of the six transcriptomes obtained result in 12,219 genes, recovered from a constructed reference transcriptome. Functional classification performed with DAVID retrieved information for 11,412 of the 12,219 annotated genes, with twenty-four GO terms and 10 KEGG pathways significantly enriched. Additional analyses performed with PANTHER suggested that “Ubiquitin proteasome pathway” was significantly overrepresented and the “Cadherin signaling pathway” was subrepresented against the DAVID database for D. rerio. Taking into account the 12,219 annotated genes, 12,189 of them were found expressed by Kallisto in at least one individual, with 42 GO terms significantly enriched compared against the gene list obtained from the reference transcriptome, as well as 5 pathways from the KEGG database and 2 from PANTHER; the last gene list analysis mentioned also found 12 Protein Classes significantly overrepresented. Considering SSR markers found in specific genes, a total of 1216 primer pairs were designed that could be used in further studies in O. argentinensis and related species, and in conjunction with the 10,784 potential SNPs were identified in annotated genes, results in set of molecular markers that could be used in future mapping or population genetic studies. Taken as a whole, the results obtained in this work will be helpful to study how individuals of genus Odontesthes deals with environmental salinity changes, as well as in studies of population structure and demographic history in this species and related taxa.

De Novo Transcriptome Analysis of Plant Pathogenic Fungus Myrothecium roridum and Identification of Genes Associated with Trichothecene Mycotoxin Biosynthesis.

Myrothecium roridum is a plant pathogenic fungus that infects different crops and decreases the yield of economical crops, including soybean, cotton, corn, pepper, and tomato. Until now, the pathogenic mechanism of M. roridum has remained unclear. Different types of trichothecene mycotoxins were isolated from M. roridum, and trichothecene was considered as a plant pathogenic factor of M. roridum. In this study, the transcriptome of M. roridum in different incubation durations was sequenced using an Illumina Hiseq 2000. A total of 35,485 transcripts and 25,996 unigenes for M. roridum were obtained from 8.0 Gb clean reads. The protein–protein network of the M. roridum transcriptome indicated that the mitogen-activated protein kinases signal pathway also played an important role in the pathogenicity of M. roridum. The genes related to trichothecene biosynthesis were annotated. The expression levels of these genes were also predicted and validated through quantitative real-time polymerase chain reaction. Tri5 gene encoding trichodiene synthase was cloned and expressed, and the purified trichodiene synthase was able to catalyze farnesyl pyrophosphate into different kinds of sesquiterpenoids.Tri4 and Tri11 genes were expressed in Escherichia coli, and their corresponding enzymatic properties were characterized. The phylogenetic tree of trichodiene synthase showed a great discrepancy between the trichodiene synthase from M. roridum and other species. Our study on the genes related to trichothecene biosynthesis establishes a foundation for the M. roridum hazard prevention, thus improving the yields of economical crops.

De Novo RNA Sequencing and Transcriptome Analysis of Monascus purpureus and Analysis of Key Genes Involved in Monacolin K Biosynthesis.

Monascus purpureus is an important medicinal and edible microbial resource. To facilitate biological, biochemical, and molecular research on medicinal components of M. purpureus, we investigated the M. purpureus transcriptome by RNA sequencing (RNA-seq). An RNA-seq library was created using RNA extracted from a mixed sample of M. purpureus expressing different levels of monacolin K output. In total 29,713 unigenes were assembled from more than 60 million high-quality short reads. A BLAST search revealed hits for 21,331 unigenes in at least one of the protein or nucleotide databases used in this study. The 22,365 unigenes were categorized into 48 functional groups based on Gene Ontology classification. Owing to the economic and medicinal importance of M. purpureus, most studies on this organism have focused on the pharmacological activity of chemical components and the molecular function of genes involved in their biogenesis. In this study, we performed quantitative real-time PCR to detect the expression of genes related to monacolin K (mokA-mokI) at different phases (2, 5, 8, and 12 days) of M. purpureus M1 and M1-36. Our study found that mokF modulates monacolin K biogenesis in M. purpureus. Nine genes were suggested to be associated with the monacolin K biosynthesis. Studies on these genes could provide useful information on secondary metabolic processes in M. purpureus. These results indicate a detailed resource through genetic engineering of monacolin K biosynthesis in M. purpureus and related species.

De Novo Assembly and Transcriptome Analysis of Bulb Onion (Allium cepa L.) during Cold Acclimation Using Contrasting Genotypes.

Bulb onion (Allium cepa) is the second most widely cultivated and consumed vegetable crop in the world. During winter, cold injury can limit the production of bulb onion. Genomic resources available for bulb onion are still very limited. To date, no studies on heritably durable cold and freezing tolerance have been carried out in bulb onion genotypes. We applied high-throughput sequencing technology to cold (2°C), freezing (-5 and -15°C), and control (25°C)-treated samples of cold tolerant (CT) and cold susceptible (CS) genotypes of A. cepa lines. A total of 452 million paired-end reads were de novo assembled into 54,047 genes with an average length of 1,331 bp. Based on similarity searches, these genes were aligned with entries in the public non-redundant (nr) database, as well as KEGG and COG database. Differentially expressed genes (DEGs) were identified using log10 values with the FPKM method. Among 5,167DEGs, 491 genes were differentially expressed at freezing temperature compared to the control temperature in both CT and CS libraries. The DEG results were validated with qRT-PCR. We performed GO and KEGG pathway enrichment analyses of all DEGs and iPath interactive analysis found 31 pathways including those related to metabolism of carbohydrate, nucleotide, energy, cofactors and vitamins, other amino acids and xenobiotics biodegradation. Furthermore, a large number of molecular markers were identified from the assembled genes, including simple sequence repeats (SSRs) 4,437 and SNP substitutions of transition and transversion types of CT and CS. Our study is the first to provide a transcriptome sequence resource for Allium spp. with regard to cold and freezing stress. We identified a large set of genes and determined their DEG profiles under cold and freezing conditions using two different genotypes. These data represent a valuable resource for genetic and genomic studies of Allium spp.

De Novo Transcriptome Analysis for Kentucky Bluegrass Dwarf Mutants Induced by Space Mutation.

Kentucky bluegrass (Poa pratensis L.) is a major cool-season turfgrass requiring frequent mowing. Utilization of cultivars with slow growth is a promising method to decrease mowing frequency. In this study, two dwarf mutant selections of Kentucky bluegrass (A12 and A16) induced by space mutation were analyzed for the differentially expressed genes compared with the wild type (WT) by the high-throughput RNA-Seq technology. 253,909 unigenes were obtained by de novo assembly. 24.20% of the unigenes had a significant level of amino acid sequence identity to Brachypodium distachyon proteins, followed by Hordeum vulgare with 18.72% among the non-redundant (NR) Blastx top hits. Assembled unigenes were associated with 32 pathways using KEGG orthology terms and their respective KEGG maps. Between WT and A16 libraries, 4,203 differentially expressed genes (DEGs) were identified, whereas there were 883 DEGs between WT and A12 libraries. Further investigation revealed that the DEG pathways were mainly involved in terpenoid biosynthesis and plant hormone metabolism, which might account for the differences of plant height and leaf blade color between dwarf mutant and WT plants. Our study presents the first comprehensive transcriptomic data and gene function analysis of Poa pratensis L., providing a valuable resource for future studies in plant dwarfing breeding and comparative genome analysis for Pooideae plants.

De Novo Transcriptome and Expression Profile Analysis to Reveal Genes and Pathways Potentially Involved in Cantharidin Biosynthesis in the Blister Beetle Mylabris cichorii.

The dried body of Mylabris cichorii is well-known Chinese traditional medicine. The sesquiterpenoid cantharidin, which is secreted mostly by adult male beetles, has recently been used as an anti-cancer drug. However, little is known about the mechanisms of cantharidin biosynthesis. Furthermore, there is currently no genomic or transcriptomic information for M. cichorii. In this study, we performed de novo assembly transcriptome of M. cichorii using the Illumina Hiseq2000. A single run produced 9.19 Gb of clean nucleotides comprising 29,247 sequences, including 23,739 annotated sequences (about 81%). We also constructed two expression profile libraries (20–25 day-old adult males and 20–25 day-old adult females) and discovered 2,465 significantly differentially-expressed genes. Putative genes and pathways involved in the biosynthesis of cantharidin were then characterized. We also found that cantharidin biosynthesis in M. cichorii might only occur via the mevalonate (MVA) pathway, not via the methylerythritol 4-phosphate/deoxyxylulose 5-phosphate (MEP/DOXP) pathway or a mixture of these. Besides, we considered that cantharidin biosynthesis might be related to the juvenile hormone (JH) biosynthesis or degradation. The results of transcriptome and expression profiling analysis provide a comprehensive sequence resource for M. cichorii that could facilitate the in-depth study of candidate genes and pathways involved in cantharidin biosynthesis, and may thus help to improve our understanding of the mechanisms of cantharidin biosynthesis in blister beetles.

De Novo Transcriptome Analysis of Warburgia ugandensis to Identify Genes Involved in Terpenoids and Unsaturated Fatty Acids Biosynthesis.

The bark of Warburgia ugandensis (Canellaceae family) has been used as a medicinal source for a long history in many African countries. The presence of diverse terpenoids and abundant polyunsaturated fatty acids (PUFAs) in this organ contributes to its broad range of pharmacological properties. Despite its medicinal and economic importance, the knowledge on the biosynthesis of terpenoid and unsaturated fatty acid in W. ugandensis bark remains largely unknown. Therefore, it is necessary to construct a genomic and/or transcriptomic database for the functional genomics study on W. ugandensis. The chemical profiles of terpenoids and fatty acids between the bark and leaves of W. ugandensis were compared by gas chromatography-mass spectrometry (GC-MS) analysis. Meanwhile, the transcriptome database derived from both tissues was created using Illumina sequencing technology. In total, about 17.1 G clean nucleotides were obtained, and de novo assembled into 72,591 unigenes, of which about 38.06% can be aligned to the NCBI non-redundant protein database. Many candidate genes in the biosynthetic pathways of terpenoids and unsaturated fatty acids were identified, including 14 unigenes for terpene synthases. Furthermore, 2,324 unigenes were discovered to be differentially expressed between both tissues; the functions of those differentially expressed genes (DEGs) were predicted by gene ontology enrichment and metabolic pathway enrichment analyses. In addition, the expression of 12 DEGs with putative roles in terpenoid and unsaturated fatty acid metabolic pathways was confirmed by qRT-PCRs, which was consistent with the data of the RNA-sequencing. In conclusion, we constructed a comprehensive transcriptome dataset derived from the bark and leaf of W. ugandensis, which forms the basis for functional genomics studies on this plant species. Particularly, the comparative analysis of the transcriptome data between the bark and leaf will provide critical clues to reveal the regulatory mechanisms underlying the biosynthesis of terpenoids and PUFAs in W. ugandensis.

De-novo transcriptome analysis of the banana shrimp (Fenneropenaeus merguiensis) and identification of genes associated with reproduction and development

The banana shrimp Fenneropenaeus merguiensis is a commercially important marine crustacean for world aquaculture and fisheries. Despite this, limited genetic information is available for it and many other penaeid shrimp species. Here we present the first in-depth analysis of the transcriptional content of 8 different tissues from the banana shrimp using RNA-Seq technologies. A total of over 1 million single-end and over 49 million paired-end reads were obtained from Roche 454FLX and illumina sequencing platforms, respectively, resulting in an assembly of 124,631 transcripts with an N50 of 1,332 and mean length of 514nt. A total of 59,179 putative protein sequences obtained from the assembled transcripts were annotated using public protein sequence databases and assigned 20,430 BLAST hits, 16,866 GO terms and 13,304 KOG categories. Further analysis revealed a rich set of transcript sequences exhibiting homology with genes associated with reproduction, sex determination and development and distinguished the tissues responsible for this expression. This report adds a substantial contribution to the sequence data currently available for F. merguiensis, providing valuable resources for further research.

De Novo Assembly and Transcriptome Analysis of Wheat with Male Sterility Induced by the Chemical Hybridizing Agent SQ-1.

Wheat (Triticum aestivum L.), one of the world’s most important food crops, is a strictly autogamous (self-pollinating) species with exclusively perfect flowers. Male sterility induced by chemical hybridizing agents has increasingly attracted attention as a tool for hybrid seed production in wheat; however, the molecular mechanisms of male sterility induced by the agent SQ-1 remain poorly understood due to limited whole transcriptome data. Therefore, a comparative analysis of wheat anther transcriptomes for male fertile wheat and SQ-1–induced male sterile wheat was carried out using next-generation sequencing technology. In all, 42,634,123 sequence reads were generated and were assembled into 82,356 high-quality unigenes with an average length of 724 bp. Of these, 1,088 unigenes were significantly differentially expressed in the fertile and sterile wheat anthers, including 643 up-regulated unigenes and 445 down-regulated unigenes. The differentially expressed unigenes with functional annotations were mapped onto 60 pathways using the Kyoto Encyclopedia of Genes and Genomes database. They were mainly involved in coding for the components of ribosomes, photosynthesis, respiration, purine and pyrimidine metabolism, amino acid metabolism, glutathione metabolism, RNA transport and signal transduction, reactive oxygen species metabolism, mRNA surveillance pathways, protein processing in the endoplasmic reticulum, protein export, and ubiquitin-mediated proteolysis. This study is the first to provide a systematic overview comparing wheat anther transcriptomes of male fertile wheat with those of SQ-1–induced male sterile wheat and is a valuable source of data for future research in SQ-1–induced wheat male sterility.

De Novo transcriptome analysis of Oncomelania hupensis after molluscicide treatment by next-generation sequencing: implications for biology and future snail interventions.

The freshwater snail Oncomelania hupensis is the only intermediate host of Schistosoma japonicum, which causes schistosomiasis. This disease is endemic in the Far East, especially in mainland China. Because niclosamide is the only molluscicide recommended by the World Health Organization, 50% wettable powder of niclosamide ethanolamine salt (WPN), the only chemical molluscicide available in China, has been widely used as the main snail control method for over two decades. Recently, a novel molluscicide derived from niclosamide, the salt of quinoid-2',5-dichloro-4'-nitro-salicylanilide (Liu Dai Shui Yang An, LDS), has been developed and proven to have the same molluscicidal effect as WPN, with lower cost and significantly lower toxicity to fish than WPN. The mechanism by which these molluscicides cause snail death is not known. Here, we report the next-generation transcriptome sequencing of O. hupensis; 145,008,667 clean reads were generated and assembled into 254,286 unigenes. Using GO and KEGG databases, 14,860 unigenes were assigned GO annotations and 4,686 unigenes were mapped to 250 KEGG pathways. Many sequences involved in key processes associated with biological regulation and innate immunity have been identified. After the snails were exposed to LDS and WPN, 254 unigenes showed significant differential expression. These genes were shown to be involved in cell structure defects and the inhibition of neurohumoral transmission and energy metabolism, which may cause snail death. Gene expression patterns differed after exposure to LDS and WPN, and these differences must be elucidated by the identification and annotation of these unknown unigenes. We believe that this first large-scale transcriptome dataset for O. hupensis will provide an opportunity for the in-depth analysis of this biomedically important freshwater snail at the molecular level and accelerate studies of the O. hupensis genome. The data elucidating the molluscicidal mechanism will be of great benefit in future snail control efforts.