NCBI Non-redundant Protein Research Articles

Characterizing the Leaf Transcriptome of Chrysanthemum rhombifolium (Ling et C. Shih), a Drought Resistant, Endemic Plant From China.

Chrysanthemum rhombifolium (Ling et C. Shih), an endemic plant that is extremely well-adapted to harsh environments. However, little is known about its molecular biology of the plant's resistant traits against stress, or even its molecular biology of overall plant. To investigate the molecular biology of C. rhombifolium and mechanism of stress adaptation, we performed transcriptome sequencing of its leaves using an Illumina platform. A total of 130,891 unigenes were obtained, and 97,496 (~74.5%) unigenes were annotated in the public protein database. The similarity search indicated that 40,878 and 74,084 unigenes showed significant similarities to known proteins from NCBI non-redundant and Swissprot protein databases, respectively. Of these, 56,213 and 42,005 unigenes were assigned to the Gene Ontology (GO) database and Cluster of Orthologous Groups (COG), respectively, and 38,918 unigenes were mapped into five main categories, including 18 KEGG pathways. Metabolism was the largest category (23,128, 59.4%) among the main KEGG categories, suggesting active metabolic processes in C. rhombifolium. About 2,459 unigenes were annotated to have a role in defense mechanism or stress tolerance. Transcriptome analysis of C. rhombifolium revealed the presence of 12,925 microsatellites in 10,524 unigenes and mono, trip, and dinucleotides having higher polymorphism rates. The phylogenetic analysis based on GME gene among related species confirmed the reliability of the transcriptomic data. This work is the first genetic study of C. rhombifolium as a new plant resource of stress-tolerant genes. This large number of transcriptome sequences enabled us to comprehensively understand the basic genetics of C. rhombifolium and discover novel genes that will be helpful in the molecular improvement of chrysanthemums.

Main Pathways and Ion Channels Differentially Expressed in the Transcriptome of Male and Female Adult Angiostrongylus cantonensis Using a Deep Sequencing Approach.

Background:The adult stage is an important period in the life cycle of Angiostrongylus cantonensis, as it is at this stage that male and female worms produce thousands of fertilized eggs daily.Methods:To explore the transcriptional details of adult male and female A. cantonensis, three groups of male and female adult worms were collected, and their transcriptome profiles were analyzed using an Illumina next-generation sequencing platform. A total of 283,910,174 clean reads were obtained, and 137,626 unigenes and 237,059 transcripts were then generated. Unigenes were successfully annotated by querying the Gene Ontology (GO), the Kyoto Encyclopedia of Genes and Genomes (KEGG), NCBI non-redundant protein sequences (NR), PFAM, STRING, and SWISS-PROT databases. Then, differentially expressed genes (DEGs) between the 2 genders were identified. The GO and KEGG databases were used for DEG annotation, and a number of DEG annotations were enriched.Results:The results obtained from querying DEGs using the GO and KEGG databases revealed that male and female adult worms exhibited differences in metabolism and production. Protein phosphorylation, ion transport, and calcium transport were all significantly enriched according to GO annotation. A number of other pathways were also enriched according to KEGG enrichment annotation, including the pentose phosphate pathway, nitrogen metabolism, oocyte meiosis pathway, neuroactive ligand-receptor interaction, calcium signaling pathway, transforming growth factor β (TGF-β) signaling pathway etc.Conclusion:We hypothesized that the nervous system of the worm plays a key role in the physiological regulation of adult A. cantonensis, and based on this, the function of the calcium-signaling pathway should be investigated.

Identification of putative miRNAs from expressed sequence tags of Gnetum gnemon L. and their cross-kingdom targets.

Wild edible plants are often found to be rich sources of nutrients and medicinally beneficial compounds with pharmacological activities. Gnetum gnemon is a nutritionally important plant and a popular food source in parts of Assam and North-East India. Various microRNAs (miRNAs) have been recently identified in many plants; however, there are no records of identification of miRNAs in any species of Gnetum. The prediction of miRNA-target associations in G. gnemon is an important step to facilitate functional genomics studies in this species. In the present study, all known miRNAs from plants available in public domain were used to search for the conserved G. gnemon miRNA homologues in publicly accessible expressed sequence tags (ESTs) in NCBI database. An aggregate of 20 new potential miRNAs belonging to two diverse miRNA families (miR399 and miR5021) were identified through a homology-based search by following stringent filtering criteria. To investigate the potential cross-kingdom effects of the identified miRNAs, we further identified the putative target genes of G. gnemon miRNAs in human transcriptome and analyzed them against the NCBI non-redundant protein database. The KEGG analysis of the target genes indicated that these genes were involved in different metabolic pathways such as caffeine metabolism, drug metabolism, and nitrotoluene degradation. The target genes of G. gnemon miRNAs in humans were found to be associated with various disorders of both hereditary and non-hereditary origin. These results could help to shed new light on understanding of miRNA-mRNAs functional networks in this species and its potential use as a small RNA-based therapy against some human diseases.

Comparative transcriptome analysis of three gonadal development stages reveals potential genes involved in gametogenesis of the fluted giant clam (Tridacna squamosa)

BackgroundGonad development and differentiation is an essential function for all sexually reproducing species, and many aspects of these developmental processes are highly conserved among the metazoa. However, the mechanisms underlying gonad development and gametogenesis remain unclear in Tridacna squamosa, a large-size bivalve of great ecological value. They are protandrous simultaneous hermaphrodites, with the male gonad maturing first, eventually followed by the female gonads. In this study, nine gonad libraries representing resting, male and hermaphrodite stages in T. squamosa were performed to identify the molecular mechanisms.ResultsSixteen thousand four hundred ninety-one unigenes were annotated in the NCBI non-redundant protein database. Among the annotated unigenes, 5091 and 7328 unigenes were assigned to Gene Ontology categories and the Kyoto Encyclopedia of Genes and Genomes (KEGG) Pathway database, respectively. A total of 4763 differentially expressed genes (DEGs) were identified by comparing male to resting gonads, consisting of 3499 which were comparatively upregulated in males and 1264 which were downregulated in males. Six hundred-ninteen DEGs between male and hermaphroditic gonads were identified, with 518 DEGs more strongly expressed in hermaphrodites and 101 more strongly expressed in males. GO (Gene Ontology) and KEGG pathway analyses revealed that various biological functions and processes, including functions related to the endocrine system, oocyte meiosis, carbon metabolism, and the cell cycle, were involved in regulating gonadal development and gametogenesis in T. squamosa. Testis-specific serine/threonine kinases 1 (TSSK1), TSSK4, TSSK5, Doublesex- and mab-3-related transcription factor 1 (DMRT1), SOX, Sperm surface protein 17 (SP17) and other genes were involved in male gonadal development in Tridacna squamosal. Both spermatogenesis- (TSSK4, spermatogenesis-associated protein 17, spermatogenesis-associated protein 8, sperm motility kinase X, SP17) and oogenesis-related genes (zona pellucida protein, Forkhead Box L2, Vitellogenin, Vitellogenin receptor, 5-hydroxytryptamine, 5-hydroxytryptamine receptor) were simultaneously highly expressed in the hermaphroditic gonad to maintain the hermaphroditism of T. squamosa.ConclusionAll these results from our study will facilitate better understanding of the molecular mechanisms underlying giant clam gonad development and gametogenesis, which can provided a base on obtaining excellent gametes during the seed production process for giant clams.

TOA: A software package for automated functional annotation in non-model plant species.

The increase of sequencing capacity provided by high-throughput platforms has made it possible to routinely obtain large sets of genomic and transcriptomic sequences from model and non-model organisms. Subsequent genomic analysis and gene discovery in next-generation sequencing experiments are, however, bottlenecked by functional annotation. One common way to perform functional annotation of sets of sequences obtained from next-generation sequencing experiments, is by searching for homologous sequences and accessing the related functional information deposited in genomic databases. Functional annotation is especially challenging for non-model organisms, like many plant species. In such cases, existing free and commercial general-purpose applications may not offer complete and accurate results. We present TOA (Taxonomy-oriented annotation), a Python-based user-friendly open source application designed to establish functional annotation pipelines geared towards non-model plant species that can run in Linux/Mac computers, HPCs and cloud servers. TOA performs homology searches against proteins stored in the PLAZA databases, NCBI RefSeq Plant, Nucleotide Database and Non-Redundant Protein Sequence Database, and outputs functional information from several ontology systems: Gene Ontology, InterPro, EC, KEGG, Mapman and MetaCyc. The software performance was validated by comparing the runtimes, total number of annotated sequences and accuracy of the functional information obtained for several plant benchmark data sets with TOA and other functional annotation solutions. TOA outperformed the other software in terms of number of annotated sequences and accuracy of the annotation and constitutes a good alternative to improve functional annotation in plants. TOA is especially recommended for gymnosperms or for low quality sequence data sets of non-model plants.

Comparative transcriptome analysis of Sweetpotato (Ipomoea batatas L.) and discovery of genes involved in starch biosynthesis

Sweetpotato, Ipomoea batatas L., Convolvulaceae, rich in starch, is a major staple economically important root crop worldwide. Limited functional genomics resources and whole genome studies could be improved through the development of next-generation sequencing technologies. Illumina sequencing of three sweetpotato varieties, Xushu 18, Xu 781 and Xushu 25, including de novo transcriptome assembly, functional annotation, and differential expression of potential starch biosynthesis-related genes are described. A total of 2508, 1807 and 3169 Mb clean reads were generated for Xushu 18, Xu 781 and Xushu 25, and were assembled into 32,112, 35,788 and 36,955 unigenes with an average length of 787, 802 and 824 bp, respectively. BLAST against the NCBI non-redundant protein, the Kyoto Encyclopedia of Genes and Genomes (KEGG) and the Gene Ontology (GO) databases have classified these unigenes into functional categories for understanding gene functions and regulation pathways. The proportion of up-regulated DEGs was lower than that of the down-regulated DEGs among three sweetpotato cultivars. The similar proportion of the up/down-regulated gene number was presented between Xushu 25 and one of its parents (Xushu18 and Xu 781). Moreover, 14 unigenes related to starch biosynthesis in Xushu 18 and Xu 781were searched out from the transcriptome sequencing data. Furthermore, gene expression of these genes in storage root during different developing stages was also investigated, 6 types of gene expression patterns were summarized. The novel sequence dataset developed in this study will be helpful for the further genetic characterization and studies of starch biosynthesis in sweetpotato.

Comparative transcriptome analysis reveals heat stress-responsive genes and their signalling pathways in lilies (Lilium longiflorum vs. Lilium distichum)

The lily, a famous bulbous flower, is seriously affected by high temperatures, which affect their growth and production. To date, the signalling pathways and the molecular mechanisms related to heat response in Lilium have not been elucidated. In this study, a comparative transcriptome analysis was performed in an important thermo-tolerant flower, L. longiflorum, and a thermo-sensitive flower, L. distichum. Lily seedlings were first exposed to heat stress at 42°C for different lengths of time, and the optimal time-points (2 h and 24 h) were selected for RNA sequencing (RNA-seq). Approximately 66.51, 66.21, and 65.36 Mb clean reads were identified from three libraries of L. longiflorum (LL_CK, LL_T2h and LL_T24h, respectively) and 66.18, 66.03, and 65.16 Mb clean reads were obtained from three libraries of L. distichum (LD_CK, LD_T2h and LD_T24h, respectively) after rRNA removing. A total of 34,301 unigenes showed similarity to known proteins in the database NCBI non-redundant protein (NR), Swiss-Prot proteins, InterPro proteins, Clusters of Orthologous Groups (COG) and Kyoto Encyclopedia of Genes and Genomes (KEGG). In addition, 1,621 genes were differentially expressed in the overlapping libraries between LL_DEGs and LD_DEGs; of these genes, 352 DEGs were obviously upregulated in L. longiflorum and downregulated in L. distichum during heat stress, including 4-coumarate, CoA ligase (4CL), caffeoyl-CoA O-methyltransferase (CCoAOMT), peroxidase, pathogenesis-related protein 10 family genes (PR10s), 14-3-3 protein, leucine-rich repeat receptor-like protein kinase, and glycine-rich cell wall structural protein-like. These genes were mainly involved in metabolic pathways, phenylpropanoid biosynthesis, plant-pathogen interactions, plant hormone signal transduction, and kinase signalling pathways. Quantitative RT-PCR was performed to validate the expression profiling of these DEGs in RNA-seq data. Taken together, the results obtained in the present study provide a comprehensive sequence resource for the discovery of heat-resistance genes and reveal potential key components that are responsive to heat stress in lilies, which may help to elucidate the heat signal transcription networks and facilitate heat-resistance breeding in lily.

Integrative Transcriptome and Proteome Analysis Identifies Major Molecular Regulation Pathways Involved in Ramie (Boehmeria nivea (L.) Gaudich) under Nitrogen and Water Co-Limitation.

Water and N are the most important factors affecting ramie (Boehmeria nivea (L.) Gaudich) growth. In this study, de novo transcriptome assembly and Tandem Mass Tags (TMT) based quantitative proteome analysis of ramie under nitrogen and water co-limitation conditions were performed, and exposed to treatments, including drought and N-deficit (WdNd), proper water but N-deficit (WNd), proper N but drought (WdN), and proper N and water (CK), respectively. A total of 64,848 unigenes (41.92% of total unigenes) were annotated in at least one database, including NCBI non-redundant protein sequences (Nr), Swiss-Prot, Protein family (Pfam), Gene Ontology (GO) and KEGG Orthology (KO), and 4268 protein groups were identified. Most significant changes in transcript levels happened under water-limited conditions, but most significant changes in protein level happened under water-limited conditions only with proper N. Poor correlation between differentially expressed genes (DEGs) and differentially expressed proteins (DEPs) was observed in ramie responding to the treatments. DEG/DEP regulation patterns related to major metabolic processes responding to water and N deficiency were analyzed, including photosynthesis, ethylene responding, glycolysis, and nitrogen metabolism. Moreover, 41 DEGs and 61 DEPs involved in regulating adaptation of ramie under water and N stresses were provided in the study, including DEGs/DEPs related to UDP—glucuronosyhransferase (UGT), ATP synthase, and carbonate dehydratase. The strong dependency of N-response of ramie on water conditions at the gene and protein levels was highlighted. Advices for simultaneously improving water and N efficiency in ramie were also provided, especially in breeding N efficient varieties with drought resistance. This study provided extensive new information on the transcriptome, proteome, their correlation, and diversification in ramie responding to water and N co-limitation.

Transcriptome analysis of field-grown Asian ginseng provides clues to environmental conditions and developmental mechanisms related to red skin root syndrome

Red skin root greatly decreases the quality and production of Asian ginseng (Panax ginseng). However, the molecular mechanisms of its origin and development are still obscure. The present study used Illumina high-throughput mRNA sequencing (RNA-seq) technology to conduct a comparative transcriptome analysis on field-sampled healthy and red skin root-affected ginseng fibrous roots. A total of 116,537 transcripts were assembled with an average length of 794 bp. Results of BLAST searches in the NCBI non-redundant protein database showed that the annotated transcripts were mostly enriched in plant species as well as the arbuscular mycorrhizae fungus Rhizophagus irregularis and the putative fungal pathogen Marssonina brunnea. With a cutoff of 1.5-fold (p<0.01), the fibrous roots of red skin root-affected ginseng showed 1856 up-regulated genes and 1581 down-regulated genes compared with the healthy ginseng roots. In addition to Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enriched analyses, the environmental stimuli and phenolic compound metabolism underlying the origin and development of red skin disease were also discussed. The present study provides molecular clues to understanding the occurrence mechanisms of red skin root in ginseng.

Description of Virulent Factors and Horizontal Gene Transfers of Keratitis-Associated Amoeba Acanthamoeba Triangularis by Genome Analysis.

Acanthamoeba triangularis strain SH 621 is a free-living amoeba belonging to Acanthamoeba ribo-genotype T4. This ubiquitous protist is among the free-living amoebas responsible for Acanthamoeba keratitis, a severe infection of human cornea. Genome sequencing and genomic comparison were carried out to explore the biological functions and to better understand the virulence mechanism related to the pathogenicity of Acanthamoeba keratitis. The genome assembly harbored a length of 66.43 Mb encompassing 13,849 scaffolds. The analysis of predicted proteins reported the presence of 37,062 ORFs. A complete annotation revealed 33,168 and 16,605 genes that matched with NCBI non-redundant protein sequence (nr) and Cluster of Orthologous Group of proteins (COG) databases, respectively. The Kyoto Encyclopedia of Genes and Genomes Pathway (KEGG) annotation reported a great number of genes related to carbohydrate, amino acid and lipid metabolic pathways. The pangenome performed with 8 available amoeba genomes belonging to genus Acanthamoeba revealed a core genome containing 843 clusters of orthologous genes with a ratio core genome/pangenome of less than 0.02. We detected 48 genes related to virulent factors of Acanthamoeba keratitis. Best hit analyses in nr database identified 99 homologous genes shared with amoeba-resisting microorganisms. This study allows the deciphering the genome of a free-living amoeba with medical interest and provides genomic data to better understand virulence-related Acanthamoeba keratitis.

Next-Generation Sequencing Reveals That Oxidative Phosphorylation Might Be a Key Pathway Differently Expressed in the Third and Fourth Stages Larvae of Angiostrongylus cantonensis.

Background:When Angiostrongylus cantonensis develops from the third and fourth stage, it needs to change its host from the middle host, snail to the final host, rat. However, the mechanism involved in this change remains unclear.Methods:The transcriptome differences of the third and fourth stages of A. cantonensis were explored by next-generation Illumina Hiseq/Miseq sequencing in China, in 2018.Results:Overall, 137 956 488 clean reads and 20 406 213 373 clean bases of the two stages larvae were produced. Based on the queries against the Gene Ontology (GO), NCBI non-redundant protein sequences (Nr), Swissprot, and Kyoto Encyclopedia of Genes and Genomes (KEGG) databases, 14 204 differentially expressed genes (DEGs) were predicted. GO enrichment analysis revealed 5660 DEGs with the top s categories as followings: biological process (GO:0008150, related to 5345 DEGs), cellular component (GO:0005575, related to 5297 DEGs), molecular function (GO:0003674, related to 5290 DEGs). In KEGG enrichment analysis, 116 genes were related to oxidative phosphorylation and 49 genes involved in the glycolytic process.Conclusion:Metabolism changes, especially oxidative phosphorylation and glycolysis, might play a key role in A. cantonensis infection of its final rat host. Many other pathways might also contribute to the transcriptome changes between these two life stages. Overall, additional studies are needed for further details.

De novo transcriptome analysis and identification of genes associated with immunity, detoxification and energy metabolism from the fat body of the tephritid gall fly, Procecidochares utilis.

The fat body, a multifunctional organ analogous to the liver and fat tissue of vertebrates, plays an important role in insect life cycles. The fat body is involved in protein storage, energy metabolism, elimination of xenobiotics, and production of immunity regulator-like proteins. However, the molecular mechanism of the fat body’s physiological functions in the tephritid stem gall-forming fly, Procecidochares utilis, are still unknown. In this study, we performed transcriptome analysis of the fat body of P. utilis using Illumina sequencing technology. In total, 3.71 G of clean reads were obtained and assembled into 30,559 unigenes, with an average length of 539 bp. Among those unigenes, 21,439 (70.16%) were annotated based on sequence similarity to proteins in NCBI’s non-redundant protein sequence database (Nr). Sequences were also compared to NCBI’s non-redundant nucleotide sequence database (Nt), a manually curated and reviewed protein sequence database (SwissProt), and KEGG and gene ontology annotations were applied to better understand the functions of these unigenes. A comparative analysis was performed to identify unigenes related to detoxification, immunity and energy metabolism. Many unigenes involved in detoxification were identified, including 50 unigenes of putative cytochrome P450s (P450s), 18 of glutathione S-transferases (GSTs), 35 of carboxylesterases (CarEs) and 26 of ATP-binding cassette (ABC) transporters. Many unigenes related to immunity were identified, including 17 putative serpin genes, five peptidoglycan recognition proteins (PGRPs) and four lysozyme genes. In addition, unigenes potentially involved in energy metabolism, including 18 lipase genes, five fatty acid synthase (FAS) genes and six elongases of very long chain fatty acid (ELOVL) genes, were identified. This transcriptome improves our genetic understanding of P. utilis and the identification of a numerous transcripts in the fat body of P. utilis offer a series of valuable molecular resources for future studies on the functions of these genes.

Transcriptomic Revelation of Phenolic Compounds Involved in Aluminum Toxicity Responses in Roots of Cunninghamia lanceolata (Lamb.) Hook.

Chinese fir (Cunninghamia lanceolata (Lamb.) Hook.) is one of the most important coniferous evergreen tree species in South China due to its desirable attributes of fast growth and production of strong and hardy wood. However, the yield of Chinese fir is often inhibited by aluminum (Al) toxicity in acidic soils of South China. Understanding the molecular mechanisms of Chinese fir root responses to Al toxicity might help to further increase its productivity. Here we used the Illumina Hiseq4000 platform to carry out transcriptome analysis of Chinese fir roots subjected to Al toxicity conditions. A total of 88.88 Gb of clean data was generated from 12 samples and assembled into 105,732 distinct unigenes. The average length and N50 length of these unigenes were 839 bp and 1411 bp, respectively. Among them, 58362 unigenes were annotated through searches of five public databases (Nr: NCBI non-redundant protein sequences, Swiss-Prot: A manually annotated and reviewed protein sequence database, GO: Gene Ontology, KOG/COG: Clusters of Orthologous Groups of proteins, and KEGG: the Kyoto Encyclopedia of Genes and Genomes database), which led to association of unigenes with 44 GO terms. Plus, 1615 transcription factors (TFs) were functionally classified. Then, differentially expressed genes (DEGs, |log2(fold change)| ≥ 1 and FDR ≤ 0.05) were identified in comparisons labelled TC1 (CK-72 h/CK-1 h) and TC2 (Al-72 h/Al-1 h). A large number of TC2 DEGs group were identified, with most being down-regulated under Al stress, while TC1 DEGs were primarily up-regulated. Combining GO, KEGG, and MapMan pathway analysis indicated that many DEGs are involved in primary metabolism, including cell wall metabolism and lipid metabolism, while other DEGs are associated with signaling pathways and secondary metabolism, including flavonoids and phenylpropanoids metabolism. Furthermore, TFs identified in TC1 and TC2 DEGs represented 21 and 40 transcription factor families, respectively. Among them, expression of bHLH, C2H2, ERF, bZIP, GRAS, and MYB TFs changed considerably under Al stress, which suggests that these TFs might play crucial roles in Chinese fir root responses to Al toxicity. These differentially expressed TFs might act in concert with flavonoid and phenylpropanoid pathway genes in fulfilling of key roles in Chinese fir roots responding to Al toxicity.

RNA-Seq analysis of Clerodendrum inerme (L.) roots in response to salt stress

BackgroundClerodendrum inerme (L.) Gaertn, a halophyte, usually grows on coastal beaches as an important mangrove plant. The salt-tolerant mechanisms and related genes of this species that respond to short-term salinity stress are unknown for us. The de novo transcriptome of C. inerme roots was analyzed using next-generation sequencing technology to identify genes involved in salt tolerance and to better understand the response mechanisms of C. inerme to salt stress.ResultsIllumina RNA-sequencing was performed on root samples treated with 400 mM NaCl for 0 h, 6 h, 24 h, and 72 h to investigate changes in C. inerme in response to salt stress. The de novo assembly identified 98,968 unigenes. Among these unigenes, 46,085 unigenes were annotated in the NCBI non-redundant protein sequences (NR) database, 34,756 sequences in the Swiss-Prot database and 43,113 unigenes in the evolutionary genealogy of genes: Non-supervised Orthologous Groups (eggNOG) database. 52 Gene Ontology (GO) terms and 31 Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways were matched to those unigenes. Most differentially expressed genes (DEGs) related to the GO terms “single-organism process”, “membrane” and “catalytic activity” were significantly enriched while numerous DEGs related to the plant hormone signal transduction pathway were also significantly enriched. The detection of relative expression levels of 9 candidate DEGs by qRT-PCR were basically consistent with fold changes in RNA sequencing analysis, demonstrating that transcriptome data can accurately reflect the response of C. inerme roots to salt stress.ConclusionsThis work revealed that the response of C. inerme roots to saline condition included significant alteration in response of the genes related to plant hormone signaling. Besides, our findings provide numerous salt-tolerant genes for further research to improve the salt tolerance of functional plants and will enhance research on salt-tolerant mechanisms of halophytes.

De Novo RNA-seq and Functional Annotation of Haemaphysalis longicornis.

Haemaphysalis longicornis (Neumann) is a hematophagous tick widely distributed in northern China. It not only causes enormous economic loss to animal husbandry, but also as a vector and reservoir of various zoonotic pathogens, it spreads natural focal diseases, such as severe fever with thrombocytopenia syndrome, seriously threatening human health. Lack of transcriptomic and genomic data from H. longicornis limits the study of this important medical vector. The engorged female H. longicornis from Gansu, China, was used for RNA extraction, de novo RNA-seq, functional annotation, and ORF prediction. As a result, 53.09 million clean reads (98.88%) with a GC content of 54.29% were obtained. A total of 65,916 Unigenes were assembled, of which 34.59% (23,330) were successfully annotated. Of these Unigenes, 22,587 (34.27%) were annotated to species by NCBI non-redundant protein (nr). Ixodes scapularis, Limulus polyphemus, Parasteatoda tepidariorum, Stegodyphus mimosarum, and Metaseiulus occidentalis were the top BLAST hit species, accounting for 47.23%, 9.58%, 4.11%, 3.50%, and 2.69%, respectively. A total of 29,182 ORFs were predicted, and 35 complete ORFs for functional genes were identified, including ORFs involved in digestion (14), stress responses (8), anticoagulation (3), reproduction (3), antimicrobial (2), drug resistance (2), movement (2), autophagy (1), and immunity (1), respectively. The Unigene ORFs encoding cathepsin and heat shock proteins were further analyzed phylogenetically. De novo RNA-seq and functional annotation of H. longicornis were successfully completed for the first time, providing a molecular data resource for further research on blood-sucking, pathogen transmission mechanisms, and effective prevention and control strategies.

Mining cytochrome p450 genes through next generation sequencing and metagenomic analysis from Binh Chau hot spring

Cytochrome P450s (CYPs) are one of the largest distributed enzymes, which catalyze more than 20 different reactions. At present, there has been an increasing realization of the power of P450 biocatalysts for the industrial synthesis of pharmaceuticals, agrochemicals, bulk chemicals, food ingredients, etc. On the other hand, the conditions of industrial processes at high temperature, high-pressure or in chemical solvent require the enzymes, which catalyze the bioconversion, have a specific properties such as thermostability, chemical tolerance or barophilicity. Up to date, the number of thermostable P450s is limited. Nowadays, DNA-metagenome technique gives us a chance to catch novel genes and unique interesting enzymes from microbial community in certain ecology. In this paper, metagenomic DNA extracted from water samples from Binh Chau hot spring was sequenced using Illumila’s HiSeq platform and was analysed to mining putative genes encoding cytochrome P450. The sequencing generated 9.4 Gb of reads containing 156,093 putative ORFs, of these, 106,903 genes were annotated in NCBI non-redundant protein sequence database. Among all the ORFs were annotated, 68 putative ORFs encoding cytochrome P450 were found belong to 36 specific groups of cytochrome P450 protein family. Of these, the melting temperature (Tm) from thirty-six completed ORFs was predicted for a better understanding of thermodynamic stability.

Mining cytochrome p450 genes through next generation sequencing and metagenomic analysis from Binh Chau hot spring

Cytochrome P450s (CYPs) are one of the largest distributed enzymes, which catalyze more than 20 different reactions. At present, there has been an increasing realization of the power of P450 biocatalysts for the industrial synthesis of pharmaceuticals, agrochemicals, bulk chemicals, food ingredients, etc. On the other hand, the conditions of industrial processes at high temperature, high-pressure or in chemical solvent require the enzymes, which catalyze the bioconversion, have a specific properties such as thermostability, chemical tolerance or barophilicity. Up to date, the number of thermostable P450s is limited. Nowadays, DNA-metagenome technique gives us a chance to catch novel genes and unique interesting enzymes from microbial community in certain ecology. In this paper, metagenomic DNA extracted from water samples from Binh Chau hot spring was sequenced using Illumila’s HiSeq platform and was analysed to mining putative genes encoding cytochrome P450. The sequencing generated 9.4 Gb of reads containing 156,093 putative ORFs, of these, 106,903 genes were annotated in NCBI non-redundant protein sequence database. Among all the ORFs were annotated, 68 putative ORFs encoding cytochrome P450 were found belong to 36 specific groups of cytochrome P450 protein family. Of these, the melting temperature (Tm) from thirty-six completed ORFs was predicted for a better understanding of thermodynamic stability.

Identification of glutathione-S-transferase genes by transcriptome analysis in Meteorus pulchricornis (Hymenoptera: Braconidae) and their expression patterns under stress of phoxim and cypermethrin

Meteorus pulchricornis (Wesmael) (Hymenoptera: Braconidae) is a preponderant endoparasitoid wasp, attacking the larvae of many lepidopteran pests. We present the first body transcriptome dataset for M. pulchricornis. In total, 50,781,796 clean reads were obtained and 33,144 unigenes were assembled; 15,458 unigenes showed a significant similarity (E value < 10−5) to known proteins in the NCBI non-redundant protein database. Gene ontology and cluster of orthologous group analyses were performed to classify the functions of genes. To better understand the role of glutathione-S-transferases (GSTs) in detoxification mechanism in M. pulchricornis, we identified seventeen GST genes (MpulGSTs) from the body transcriptome. Among these, fifteen MpulGSTs belonged to cytosolic GSTs and the other two belonged to microsomal classes. The cytosolic GSTs were classified into four different clades: four in delta, three in omega, seven in sigma, and one in zeta. The expression levels of these MpulGSTs after exposure to sub-lethal concentrations of phoxim and cypermethrin were determined using real-time quantitative polymerase chain reaction: seven MpulGSTs (MpulGSTD3, MpulGSTS1, MpulGSTS2, MpulGSTS4, MpulGSTS6 MpulGSTO3, and MpulGSTmic1) and 11 MpulGSTs (MpulGSTD1, MpulGSTD2, MpulGSTD3, MpulGSTO2, MpulGSTS1, MpulGSTS2, MpulGSTS3, MpulGSTS4, MpulGSTS5, MpulGSTS7, and MpulGSTmic1) were highly expressed, respectively. These results suggested that GST genes may play a pivotal role in detoxification process in M. pulchricornis. Our findings would provide a theoretical base for elucidating insecticide susceptibility and should promote functional research on specific GST genes in parasitoid wasps.

Effector proteins of Rhizophagus proliferus: conserved protein domains may play a role in host-specific interaction with different plant species.

Arbuscular mycorrhizal (AM) fungi show high promiscuity in terms of host. Effector proteins expressed by AM fungi are found important in establishing interaction with host. However, the mechanistic underlying host-specific interactions of the fungi remain unknown. The present study aimed (i) to identify effectors encoded by Rhizophagus proliferus and (ii) to understand molecular specificity encoded in effectors for interaction with specific plant species. The effectors predicted from the whole genome sequence were annotated by homology search in NCBI non-redundant protein, Interproscan, and pathogen-host interaction (PHI) databases. In total, 416 small secreted peptides (SSPs) were predicted, which were effector peptides with presence of nuclear localization signal, small cysteine-rich, and repeat-containing proteins domains. Similar to the functionally validated SP7 effectors in Rhizophagus irregularis, two proteins (RP8598 and RP23081) were identified in R. proliferus. To understand whether interaction between SP7 and the plant target protein, ERF19, is specific in nature, we examined protein-peptide interaction using in silico molecular docking. Pairwise interaction of RP8598 and RP23081 with the ethylene-responsive factors (ERF19) coded by five different plant species (Lotus japonicus, Solanum lycopersicum, Ocimum tenuiflorum, Medicago truncatula, Diospyros kaki) was investigated. Prediction of high-quality interaction of SP7 effector with ERF19 protein expressed only by specific plant species was observed in in silico molecular docking, which may reiterate the role of effectors in host specificity. The outcomes from our study indicated that sequence precision encoded in the effector peptides of AM fungi and immunomodulatory proteins of host may regulate host specificity in these fungi.

Transcriptome analysis of Ajowan (Trachyspermum ammi L.) inflorescence

Trachyspermum ammi (L.) Sprague, commonly known as ‘Ajowan’, belongs to the family ‘Apiaceae’. Ajowan fruits yield 2–6% essential oil, and thymol is the major constituent of the oil (35–60%). T. ammi is of high medicinal value; however, the genomic resources for this medicinal plant are rare. To obtain transcript sequences of ajowan inflorescence, RNA-Seq was applied using Illumina HiSeq 2000 platform. Following de novo assembly, 68051 unigenes were produced, among which 43156 unigenes were annotated against different sequence databases. Blastx results of ajowan unigenes showed that 41950 and 29273 unigenes were hit in the NCBI non-redundant (Nr) protein and UniProt databases, respectively. Gene Ontology (GO) classification showed that 29258 unigenes were categorized into 55 GO terms, and 26002 unigenes were clustered in the Orthologous Group categories. Blastx results against the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway database indicated that 14986 unigenes were represented in 127 pathways. In addition, 8751 sequences containing simple sequence repeats (SSRs) and 1180 putative transcription factor genes were also identified. As far as we know, this repository of genomic information is the first resource currently available for transcriptome characterization, gene discovery, developing SSR molecular markers and future genomic research on ajowan.