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Transcriptogenomics identification and characterization of RNA editing sites in human primary monocytes using high-depth next generation sequencing data

Leong WM, Ripen AM, Mirsafian H, Mohamad SB, Merican AF

Genomics, 2019 07;111(4):899-905.
PMID: 29885984 DOI: 10.1016/j.ygeno.2018.05.019

High-depth next generation sequencing data provide valuable insights into the number and distribution of RNA editing events. Here, we report the RNA editing events at cellular level of human primary monocyte using high-depth whole genomic and transcriptomic sequencing data. We identified over a ten thousand putative RNA editing sites and 69% of the sites were A-to-I editing sites. The sites enriched in repetitive sequences and intronic regions. High-depth sequencing datasets revealed that 90% of the canonical sites were edited at lower frequencies (<0.7). Single and multiple human monocytes and brain tissues samples were analyzed through genome sequence independent approach. The later approach was observed to identify more editing sites. Monocytes was observed to contain more C-to-U editing sites compared to brain tissues. Our results establish comparable pipeline that can address current limitations as well as demonstrate the potential for highly sensitive detection of RNA editing events in single cell type.

Matched MeSH terms: High-Throughput Nucleotide Sequencing/methods*
Characterization of genetic sequence variation of 58 STR loci in four major population groups

Novroski NMM, King JL, Churchill JD, Seah LH, Budowle B

Forensic Sci Int Genet, 2016 11;25:214-226.
PMID: 27697609 DOI: 10.1016/j.fsigen.2016.09.007

Massively parallel sequencing (MPS) can identify sequence variation within short tandem repeat (STR) alleles as well as their nominal allele lengths that traditionally have been obtained by capillary electrophoresis. Using the MiSeq FGx Forensic Genomics System (Illumina), STRait Razor, and in-house excel workbooks, genetic variation was characterized within STR repeat and flanking regions of 27 autosomal, 7 X-chromosome and 24 Y-chromosome STR markers in 777 unrelated individuals from four population groups. Seven hundred and forty six autosomal, 227 X-chromosome, and 324 Y-chromosome STR alleles were identified by sequence compared with 357 autosomal, 107 X-chromosome, and 189 Y-chromosome STR alleles that were identified by length. Within the observed sequence variation, 227 autosomal, 156 X-chromosome, and 112 Y-chromosome novel alleles were identified and described. One hundred and seventy six autosomal, 123 X-chromosome, and 93 Y-chromosome sequence variants resided within STR repeat regions, and 86 autosomal, 39 X-chromosome, and 20 Y-chromosome variants were located in STR flanking regions. Three markers, D18S51, DXS10135, and DYS385a-b had 1, 4, and 1 alleles, respectively, which contained both a novel repeat region variant and a flanking sequence variant in the same nucleotide sequence. There were 50 markers that demonstrated a relative increase in diversity with the variant sequence alleles compared with those of traditional nominal length alleles. These population data illustrate the genetic variation that exists in the commonly used STR markers in the selected population samples and provide allele frequencies for statistical calculations related to STR profiling with MPS data.

Matched MeSH terms: High-Throughput Nucleotide Sequencing*
Connecting Proteomics to Next-Generation Sequencing: Proteogenomics and Its Current Applications in Biology

Low TY, Mohtar MA, Ang MY, Jamal R

Proteomics, 2019 05;19(10):e1800235.
PMID: 30431238 DOI: 10.1002/pmic.201800235

Understanding the relationship between genotypes and phenotypes is essential to disentangle biological mechanisms and to unravel the molecular basis of diseases. Genes and proteins are closely linked in biological systems. However, genomics and proteomics have developed separately into two distinct disciplines whereby crosstalk among scientists from the two domains is limited and this constrains the integration of both fields into a single data modality of useful information. The emerging field of proteogenomics attempts to address this by building bridges between the two disciplines. In this review, how genomics and transcriptomics data in different formats can be utilized to assist proteogenomics application is briefly discussed. Subsequently, a much larger part of this review focuses on proteogenomics research articles that are published in the last five years that answer two important questions. First, how proteogenomics can be applied to tackle biological problems is discussed, covering genome annotation and precision medicine. Second, the latest developments in analytical technologies for data acquisition and the bioinformatics tools to interpret and visualize proteogenomics data are covered.

Matched MeSH terms: High-Throughput Nucleotide Sequencing/trends*
Fulltext Population Genomics of an Anadromous Hilsa Shad Tenualosa ilisha Species across Its Diverse Migratory Habitats: Discrimination by Fine-Scale Local Adaptation

Asaduzzaman M, Igarashi Y, Wahab MA, Nahiduzzaman M, Rahman MJ, Phillips MJ, et al.

Genes (Basel), 2019 12 30;11(1).
PMID: 31905942 DOI: 10.3390/genes11010046

The migration of anadromous fish in heterogenic environments unceasingly imposes a selective pressure that results in genetic variation for local adaptation. However, discrimination of anadromous fish populations by fine-scale local adaptation is challenging because of their high rate of gene flow, highly connected divergent population, and large population size. Recent advances in next-generation sequencing (NGS) have expanded the prospects of defining the weakly structured population of anadromous fish. Therefore, we used NGS-based restriction site-associated DNA (NextRAD) techniques on 300 individuals of an anadromous Hilsa shad (Tenualosa ilisha) species, collected from nine strategic habitats, across their diverse migratory habitats, which include sea, estuary, and different freshwater rivers. The NextRAD technique successfully identified 15,453 single nucleotide polymorphism (SNP) loci. Outlier tests using the FST OutFLANK and pcadapt approaches identified 74 and 449 SNPs (49 SNPs being common), respectively, as putative adaptive loci under a divergent selection process. Our results, based on the different cluster analyses of these putatively adaptive loci, suggested that local adaptation has divided the Hilsa shad population into two genetically structured clusters, in which marine and estuarine collection sites were dominated by individuals of one genetic cluster and different riverine collection sites were dominated by individuals of another genetic cluster. The phylogenetic analysis revealed that all the riverine populations of Hilsa shad were further subdivided into the north-western riverine (turbid freshwater) and the north-eastern riverine (clear freshwater) ecotypes. Among all of the putatively adaptive loci, only 36 loci were observed to be in the coding region, and the encoded genes might be associated with important biological functions related to the local adaptation of Hilsa shad. In summary, our study provides both neutral and adaptive contexts for the observed genetic divergence of Hilsa shad and, consequently, resolves the previous inconclusive findings on their population genetic structure across their diverse migratory habitats. Moreover, the study has clearly demonstrated that NextRAD sequencing is an innovative approach to explore how dispersal and local adaptation can shape genetic divergence of non-model anadromous fish that intersect diverse migratory habitats during their life-history stages.

Matched MeSH terms: High-Throughput Nucleotide Sequencing/methods*
Fulltext Complete mitochondrial genome of Dacus vijaysegarani and phylogenetic relationships with congeners and other tephritid fruit flies (Insecta: Diptera)

Yong HS, Chua KO, Song SL, Liew YJ, Eamsobhana P, Chan KG

Mol Biol Rep, 2021 Aug;48(8):6047-6056.
PMID: 34357549 DOI: 10.1007/s11033-021-06608-2

BACKGROUND: Tephritid fruit flies of the genus Dacus are members of the tribe Dacini, subfamily Dacinae. There are some 274 species worldwide, distributed in Africa and the Asia-Pacific. To date, only five complete mitochondrial genomes (mitogenomes) of Dacus fruit flies have been published and are available in the GenBank.
METHODS AND RESULTS: In view of the lack of study on their mitogenome, we sequenced (by next generation sequencing) and annotated the complete mitogenome of D. vijaysegarani from Malaysia to determine its features and phylogenetic relationship. The whole mitogenome of D. vijaysegarani has identical gene order with the published mitogenomes of the genus Dacus, with 13 protein-coding genes, two rRNA genes, 22 tRNAs, a non-coding A + T rich control region, and intergenic spacer and overlap sequences. Phylogenetic analysis based on 15 mitochondrial genes (13 PCGs and two rRNA genes), reveals Dacus, Zeugodacus and Bactrocera forming a distinct clade. The genus Dacus forms a monophyletic group in the subclade containing also the Zeugodacus group; this Dacus-Zeugodacus subclade is distinct from the Bactrocera subclade. D. (Mellesis) vijaysegarani forms a lineage with D. (Mellesis) trimacula in the subcluster containing also the lineage of D. (Mellesis) conopsoides and D. (Callantra) longicornis. D. (Dacus) bivittatus and D. (Didacus) ciliatus form a distinct subcluster. Based on cox1 sequences, the Malaysia and Vietnam taxa of D. vijaysegarani may not be conspecific.
CONCLUSIONS: Overall, the mitochondrial genome of D. vijaysegarani provided essential molecular data that could be useful for further studies for species diagnosis, evolution and phylogeny research of other tephritid fruit flies in the future.

Matched MeSH terms: High-Throughput Nucleotide Sequencing/methods
Fulltext Multi-platform discovery of haplotype-resolved structural variation in human genomes

Chaisson MJP, Sanders AD, Zhao X, Malhotra A, Porubsky D, Rausch T, et al.

Nat Commun, 2019 04 16;10(1):1784.
PMID: 30992455 DOI: 10.1038/s41467-018-08148-z

The incomplete identification of structural variants (SVs) from whole-genome sequencing data limits studies of human genetic diversity and disease association. Here, we apply a suite of long-read, short-read, strand-specific sequencing technologies, optical mapping, and variant discovery algorithms to comprehensively analyze three trios to define the full spectrum of human genetic variation in a haplotype-resolved manner. We identify 818,054 indel variants (<50 bp) and 27,622 SVs (≥50 bp) per genome. We also discover 156 inversions per genome and 58 of the inversions intersect with the critical regions of recurrent microdeletion and microduplication syndromes. Taken together, our SV callsets represent a three to sevenfold increase in SV detection compared to most standard high-throughput sequencing studies, including those from the 1000 Genomes Project. The methods and the dataset presented serve as a gold standard for the scientific community allowing us to make recommendations for maximizing structural variation sensitivity for future genome sequencing studies.

Matched MeSH terms: High-Throughput Nucleotide Sequencing/methods
Population and performance analyses of four major populations with Illumina's FGx Forensic Genomics System

Churchill JD, Novroski NMM, King JL, Seah LH, Budowle B

Forensic Sci Int Genet, 2017 09;30:81-92.
PMID: 28651097 DOI: 10.1016/j.fsigen.2017.06.004

The MiSeq FGx Forensic Genomics System (Illumina) enables amplification and massively parallel sequencing of 59 STRs, 94 identity informative SNPs, 54 ancestry informative SNPs, and 24 phenotypic informative SNPs. Allele frequency and population statistics data were generated for the 172 SNP loci included in this panel on four major population groups (Chinese, African Americans, US Caucasians, and Southwest Hispanics). Single-locus and combined random match probability values were generated for the identity informative SNPs. The average combined STR and identity informative SNP random match probabilities (assuming independence) across all four populations were 1.75E-67 and 2.30E-71 with length-based and sequence-based STR alleles, respectively. Ancestry and phenotype predictions were obtained using the ForenSeq™ Universal Analysis System (UAS; Illumina) based on the ancestry informative and phenotype informative SNP profiles generated for each sample. Additionally, performance metrics, including profile completeness, read depth, relative locus performance, and allele coverage ratios, were evaluated and detailed for the 725 samples included in this study. While some genetic markers included in this panel performed notably better than others, performance across populations was generally consistent. The performance and population data included in this study support that accurate and reliable profiles were generated and provide valuable background information for laboratories considering internal validation studies and implementation.

Matched MeSH terms: High-Throughput Nucleotide Sequencing/instrumentation*
Fulltext Computational approach to discriminate human and mouse sequences in patient-derived tumour xenografts

Callari M, Batra AS, Batra RN, Sammut SJ, Greenwood W, Clifford H, et al.

BMC Genomics, 2018 01 05;19(1):19.
PMID: 29304755 DOI: 10.1186/s12864-017-4414-y

BACKGROUND: Patient-Derived Tumour Xenografts (PDTXs) have emerged as the pre-clinical models that best represent clinical tumour diversity and intra-tumour heterogeneity. The molecular characterization of PDTXs using High-Throughput Sequencing (HTS) is essential; however, the presence of mouse stroma is challenging for HTS data analysis. Indeed, the high homology between the two genomes results in a proportion of mouse reads being mapped as human.
RESULTS: In this study we generated Whole Exome Sequencing (WES), Reduced Representation Bisulfite Sequencing (RRBS) and RNA sequencing (RNA-seq) data from samples with known mixtures of mouse and human DNA or RNA and from a cohort of human breast cancers and their derived PDTXs. We show that using an In silico Combined human-mouse Reference Genome (ICRG) for alignment discriminates between human and mouse reads with up to 99.9% accuracy and decreases the number of false positive somatic mutations caused by misalignment by >99.9%. We also derived a model to estimate the human DNA content in independent PDTX samples. For RNA-seq and RRBS data analysis, the use of the ICRG allows dissecting computationally the transcriptome and methylome of human tumour cells and mouse stroma. In a direct comparison with previously reported approaches, our method showed similar or higher accuracy while requiring significantly less computing time.
CONCLUSIONS: The computational pipeline we describe here is a valuable tool for the molecular analysis of PDTXs as well as any other mixture of DNA or RNA species.

Matched MeSH terms: High-Throughput Nucleotide Sequencing/methods*
Fulltext Cognizance of Molecular Methods for the Generation of Mutagenic Phage Display Antibody Libraries for Affinity Maturation

Lim CC, Choong YS, Lim TS

Int J Mol Sci, 2019 Apr 15;20(8).
PMID: 30991723 DOI: 10.3390/ijms20081861

Antibodies leverage on their unique architecture to bind with an array of antigens. The strength of interaction has a direct relation to the affinity of the antibodies towards the antigen. In vivo affinity maturation is performed through multiple rounds of somatic hypermutation and selection in the germinal centre. This unique process involves intricate sequence rearrangements at the gene level via molecular mechanisms. The emergence of in vitro display technologies, mainly phage display and recombinant DNA technology, has helped revolutionize the way antibody improvements are being carried out in the laboratory. The adaptation of molecular approaches in vitro to replicate the in vivo processes has allowed for improvements in the way recombinant antibodies are designed and tuned. Combinatorial libraries, consisting of a myriad of possible antibodies, are capable of replicating the diversity of the natural human antibody repertoire. The isolation of target-specific antibodies with specific affinity characteristics can also be accomplished through modification of stringent protocols. Despite the ability to screen and select for high-affinity binders, some 'fine tuning' may be required to enhance antibody binding in terms of its affinity. This review will provide a brief account of phage display technology used for antibody generation followed by a summary of different combinatorial library characteristics. The review will focus on available strategies, which include molecular approaches, next generation sequencing, and in silico approaches used for antibody affinity maturation in both therapeutic and diagnostic applications.

Matched MeSH terms: High-Throughput Nucleotide Sequencing/methods*
Transcriptome-seq provides insights into sex-preference pattern of gene expression between testis and ovary of the crucifix crab ( Charybdis feriatus)

Zhang Y, Miao G, Fazhan H, Waiho K, Zheng H, Li S, et al.

Physiol Genomics, 2018 05 01;50(5):393-405.
PMID: 29570432 DOI: 10.1152/physiolgenomics.00016.2018

The crucifix crab, Charybdis feriatus, which mainly inhabits Indo-Pacific region, is regarded as one of the most high-potential species for domestication and incorporation into the aquaculture sector. However, the regulatory mechanisms of sex determination and differentiation of this species remain unclear. To identify candidate genes involved in sex determination and differentiation, high throughput sequencing of transcriptome from the testis and ovary of C. feriatus was performed by the Illumina platform. After removing adaptor primers, low-quality sequences and very short (<50 nt) reads, we obtained 80.9 million and 66.2 million clean reads from testis and ovary, respectively. A total of 86,433 unigenes were assembled, and ~43% (37,500 unigenes) were successfully annotated to the NR, NT, Swiss-Prot, KEGG, COG, GO databases. By comparing the testis and ovary libraries, we obtained 27,636 differentially expressed genes. Some candidate genes involved in the sex determination and differentiation of C. feriatus were identified, such as vasa, pgds, vgr, hsp90, dsx-f, fem-1, and gpr. In addition, 88,608 simple sequence repeats were obtained, and 61,929 and 77,473 single nucleotide polymorphisms from testis and ovary were detected, respectively. The transcriptome profiling was validated by quantitative real-time PCR in 30 selected genes, which showed a good consistency. The present study is the first high-throughput transcriptome sequencing of C. feriatus. These findings will be useful for future functional analysis of sex-associated genes and molecular marker-assisted selections in C. feriatus.

Matched MeSH terms: High-Throughput Nucleotide Sequencing/methods*
Fulltext Ecological and Ontogenetic Components of Larval Lake Sturgeon Gut Microbiota Assembly, Successional Dynamics, and Ecological Evaluation of Neutral Community Processes

Abdul Razak S, Scribner KT

Appl Environ Microbiol, 2020 05 05;86(10).
PMID: 32169941 DOI: 10.1128/AEM.02662-19

Gastrointestinal (GI) or gut microbiotas play essential roles in host development and physiology. These roles are influenced partly by the microbial community composition. During early developmental stages, the ecological processes underlying the assembly and successional changes in host GI community composition are influenced by numerous factors, including dispersal from the surrounding environment, age-dependent changes in the gut environment, and changes in dietary regimes. However, the relative importance of these factors to the gut microbiota is not well understood. We examined the effects of environmental (diet and water sources) and host early ontogenetic development on the diversity of and the compositional changes in the gut microbiota of a primitive teleost fish, the lake sturgeon (Acipenser fulvescens), based on massively parallel sequencing of the 16S rRNA gene. Fish larvae were raised in environments that differed in water source (stream versus filtered groundwater) and diet (supplemented versus nonsupplemented Artemia fish). We quantified the gut microbial community structure at three stages (prefeeding and 1 and 2 weeks after exogenous feeding began). The diversity declined and the community composition differed significantly among stages; however, only modest differences associated with dietary or water source treatments were documented. Many taxa present in the gut were over- or underrepresented relative to neutral expectations in each sampling period. The findings indicate dynamic relationships between the gut microbiota composition and host gastrointestinal physiology, with comparatively smaller influences being associated with the rearing environments. Neutral models of community assembly could not be rejected, but selectivity associated with microbe-host GI tract interactions through early ontogenetic stages was evident. The results have implications for sturgeon conservation and aquaculture production specifically and applications of microbe-based management in teleost fish generally.IMPORTANCE We quantified the effects of environment (diet and water sources) and host early ontogenetic development on the diversity of and compositional changes in gut microbial communities based on massively parallel sequencing of the 16S rRNA genes from the GI tracts of larval lake sturgeon (Acipenser fulvescens). The gut microbial community diversity declined and the community composition differed significantly among ontogenetic stages; however, only modest differences associated with dietary or water source treatments were documented. Selectivity associated with microbe-host GI tract interactions through early ontogenetic stages was evident. The results have implications for lake sturgeon and early larval ecology and survival in their natural habitat and for conservation and aquaculture production specifically, as well as applications of microbe-based management in teleost fish generally.

Matched MeSH terms: High-Throughput Nucleotide Sequencing/veterinary
A nucleotide signature for identification of Aglaia stellatopilosa Pannell

Ng BL, Omarzuki M, Lau GS, Pannell CM, Yeo TC

Mol Biotechnol, 2014 Jul;56(7):671-9.
PMID: 24623047 DOI: 10.1007/s12033-014-9746-0

Members of the genus Aglaia have been reported to contain bioactive phytochemicals. The genus, belonging to the Meliaceae family, is represented by at least 120 known species of woody trees or shrubs in the tropical rain forest. As some of these species are very similar in their morphology, taxonomic identification can be difficult. A reliable and definitive molecular method which can identify Aglaia to the level of the species will hence be useful in comparing the content of specific bioactive compounds between the species of this genus. Here, we report the analysis of DNA sequences in the internal transcribed spacer (ITS) of the nuclear ribosomal DNA and the observation of a unique nucleotide signature in the ITS that can be used for the identification of Aglaia stellatopilosa. The nucleotide signature consists of nine bases over the length of the ITS sequence (654 bp). This uniqueness was validated in 37 samples identified as Aglaia stellatopilosa by an expert taxonomist, whereas the nucleotide signature was lacking in a selection of other Aglaia species and non-Aglaia genera. This finding suggests that molecular typing could be utilized in the identification of Aglaia stellatopilosa.

Matched MeSH terms: High-Throughput Nucleotide Sequencing
Development of novel tetra- and trinucleotide microsatellite markers for giant grouper Epinephelus lanceolatus using 454 pyrosequencing

Kim KS, Noh CH, Moon SJ, Han SH, Bang IC

Mol Biol Rep, 2016 Jun;43(6):541-8.
PMID: 27059503 DOI: 10.1007/s11033-016-3980-4

Giant grouper (Epinephelus lanceolatus) is a commercially important species, but its wild population has recently been classified as vulnerable. This species has significant potential for use in aquaculture, though a greater understanding of population genetics is necessary for selective breeding programs to minimize kinship for genetically healthy individuals. High-throughput pyrosequencing of genomic DNA was used to identify and characterize novel tetra- and trinucleotide microsatellite markers in giant grouper from Sabah, Malaysia. In total, of 62,763 sequences containing simple sequence repeats (SSRs) were obtained, and 78 SSR loci were selected to possibly contain tetra- and trinucleotide repeats. Of these loci, 16 had tetra- and 8 had trinucleotide repeats, all of which exhibited polymorphisms within easily genotyped regions. A total of 143 alleles were identified with an average of 5.94 alleles per locus, with mean observed and expected heterozygosities of 0.648 and 0.620, respectively. Among of them, 15 microsatellite markers were identified without null alleles and with Hardy-Weinberg equilibrium. These alleles showed a combined non-exclusion probability of 0.01138. The probability of individual identification (PID) value combined with in descending order 12 microsatellite markers was 0.00008, which strongly suggests that the use of the microsatellite markers developed in this study in various combinations would result in a high resolution method for parentage analysis and individual identification. These markers could be used to establish a broodstock management program for giant grouper and to provide a foundation for genetic studies such as population structure, parentage analysis, and kinship selection.

Matched MeSH terms: High-Throughput Nucleotide Sequencing
Fulltext Finding Nemo: hybrid assembly with Oxford Nanopore and Illumina reads greatly improves the clownfish (Amphiprion ocellaris) genome assembly

Tan MH, Austin CM, Hammer MP, Lee YP, Croft LJ, Gan HM

Gigascience, 2018 03 01;7(3):1-6.
PMID: 29342277 DOI: 10.1093/gigascience/gix137

Background: Some of the most widely recognized coral reef fishes are clownfish or anemonefish, members of the family Pomacentridae (subfamily: Amphiprioninae). They are popular aquarium species due to their bright colours, adaptability to captivity, and fascinating behavior. Their breeding biology (sequential hermaphrodites) and symbiotic mutualism with sea anemones have attracted much scientific interest. Moreover, there are some curious geographic-based phenotypes that warrant investigation. Leveraging on the advancement in Nanopore long read technology, we report the first hybrid assembly of the clown anemonefish (Amphiprion ocellaris) genome utilizing Illumina and Nanopore reads, further demonstrating the substantial impact of modest long read sequencing data sets on improving genome assembly statistics.
Results: We generated 43 Gb of short Illumina reads and 9 Gb of long Nanopore reads, representing approximate genome coverage of 54× and 11×, respectively, based on the range of estimated k-mer-predicted genome sizes of between 791 and 967 Mbp. The final assembled genome is contained in 6404 scaffolds with an accumulated length of 880 Mb (96.3% BUSCO-calculated genome completeness). Compared with the Illumina-only assembly, the hybrid approach generated 94% fewer scaffolds with an 18-fold increase in N50 length (401 kb) and increased the genome completeness by an additional 16%. A total of 27 240 high-quality protein-coding genes were predicted from the clown anemonefish, 26 211 (96%) of which were annotated functionally with information from either sequence homology or protein signature searches.
Conclusions: We present the first genome of any anemonefish and demonstrate the value of low coverage (∼11×) long Nanopore read sequencing in improving both genome assembly contiguity and completeness. The near-complete assembly of the A. ocellaris genome will be an invaluable molecular resource for supporting a range of genetic, genomic, and phylogenetic studies specifically for clownfish and more generally for other related fish species of the family Pomacentridae.

Matched MeSH terms: High-Throughput Nucleotide Sequencing
Assessing diet of the rufous-winged philentoma (philentoma pyrhoptera) in lowland tropical forest using next-generation sequencing

Mohammad Saiful Mansor, Shukor Md. Nor, Rosli Ramli

Sains Malaysiana, 2018;47:1045-1050.

Dietary study provides understanding in predator-prey relationships, yet diet of tropical forest birds is poorly understood.
In this study, a non-invasive method, next-generation sequencing (Illumina MiSeq platform) was used to identify prey in
the faecal samples of the Rufous-winged Philentoma (Philentoma pyrhoptera). Dietary samples were collected in lowland
tropical forest of central Peninsular Malaysia. A general invertebrate primer pair was used for the first time to assess
diet of tropical birds. The USEARCH was used to cluster the COI mtDNA sequences into Operational Taxonomic Unit (OTU).
OTU sequences were aligned and queried through the GenBank or Biodiversity of Life Database (BOLD). We identified
26 distinct arthropod taxa from 31 OTUs. Of all OTUs, there was three that could be identified up to species level, 20 to
genus level, three to family level and five could not assigned to any taxa (the BLAST hits were poor). All sequences were
identified to class Insecta belonging to 18 families from four orders, where Lepidoptera representing major insect order
consumed by study bird species. This non-invasive molecular approach provides a practical and rapid technique to
understand of how energy flows across ecosystems. This technique could be very useful to screen for possible particular
pest insects consumed by insectivores (e.g. birds and bats) in crop plantation. A comprehensive arthropod studies and
local reference sequences need to be added to the database to improve the proportion of sequences that can be identified.

Matched MeSH terms: High-Throughput Nucleotide Sequencing
Fulltext Complete chloroplast genome sequence of Gigantochloa verticillata (Bambusodae)

Zhang YY, Fan LL, Zheng FY, Zhao T, Rong JD, Chen LG, et al.

Mitochondrial DNA B Resour, 2020 Feb 06;5(1):306-307.
PMID: 33366532 DOI: 10.1080/23802359.2019.1702484

Gigantochloa verticillata is produced in Mengla and Jinghong, Yunnan Province, China, and cultivated in Hong Kong. Vietnam, Thailand, India, Indonesia, and Malaysia are distributed and cultivated. We determined the complete chloroplast genome sequence for G. verticillata using Illumina sequencing data. The complete chloroplast sequence is 139,489 bp, including large single-copy (LSC) region of 83,062 bp, small single-copy (SSC) region of 12,877 bp, and a pair of invert repeats (IR) regions of 21,775 bp. Plastid genome contain 132 genes, 85 protein-coding genes, 39 tRNA genes, and 8 rRNA genes. Phylogenetic analysis based on 23 chloroplast genomes indicates that G. verticillata is closely related to Dendrocalamus latiflorus in Bambusodae.

Matched MeSH terms: High-Throughput Nucleotide Sequencing
Fulltext Transcriptome data of the carrageenophyte Eucheuma denticulatum

Othman R, Abd Rasib AA, Ilias MA, Murthy S, Ismail N, Mohd Hanafi N

Data Brief, 2019 Jun;24:103824.
PMID: 30984808 DOI: 10.1016/j.dib.2019.103824

Eucheuma denticulatum or commonly known as "Spinosum", is an economically important red alga that naturally grows on coral reefs with moderately strong currents in tropical and sub-tropical areas. This species is the primary source of iota-carrageenan which has high demands in the food, pharmaceutical and manufacturing industries, and as such it has been widely cultivated. The increasing global demand for carrageenan has led to extensive commercial cultivation of carrageenophytes mainly in the tropics. The carrageenophyte seaweeds including E. denticulatum are indigenous to Sabah, Malaysia. To enrich the information on the genes involved in carrageenan biosynthesis, RNA sequencing has been performed and transcriptomic dataset has been generated using Illumina HiSeq™ 2000 sequencer. The raw data and transcriptomic data have been deposited in NCBI database with the accession number PRJNA477734. These data will provide valuable resources for functional genomics annotation and investigation of mechanisms underlying the regulations of genes in this algal species.

Matched MeSH terms: High-Throughput Nucleotide Sequencing
Disruption of methicillin-resistant Staphylococcus aureus protein synthesis by tannins

Adnan SN, Ibrahim N, Yaacob WA

Germs, 2017 Dec;7(4):186-192.
PMID: 29264356 DOI: 10.18683/germs.2017.1125

Introduction: Methicillin-resistant Staphylococcus aureus (MRSA) is a worldwide public health threat, displaying multiple antibiotic resistance that causes morbidity and mortality. Management of multidrug-resistant (MDR) MRSA infections is extremely difficult due to their inherent resistance to currently used antibiotics. New antibiotics are needed to combat the emergence of antimicrobial resistance.
Methods: The in vitro effect of tannins was studied against MRSA reference strain (ATCC 43300) and MRSA clinical strains utilizing antimicrobial assays in conjunction with both scanning and transmission electron microscopy. To reveal the influence of tannins in MRSA protein synthesis disruption, we utilized next-generation sequencing (NGS) to provide further insight into the novel protein synthesis transcriptional response of MRSA exposed to these compounds.
Results: Tannins possessed both bacteriostatic and bactericidal activity with minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of 0.78 and 1.56 mg/mL, respectively, against all tested MRSA. Scanning and transmission electron microscopy of MRSA treated with tannins showed decrease in cellular volume, indicating disruption of protein synthesis.
Conclusion: Analysis of a genome-wide transcriptional profile of the reference strain ATCC 43300 MRSA in response to tannins has led to the finding that tannins induced significant modulation in essential ribosome pathways, which caused a reduction in the translation processes that lead to inhibition of protein synthesis and obviation of bacterial growth. These findings highlight the potential of tannins as new promising anti-MRSA agents in clinical application such as body wash and topical cream or ointments.

Matched MeSH terms: High-Throughput Nucleotide Sequencing
G-quadruplex structures bind to EZ-Tn5 transposase

Cree SL, Chua EW, Crowther J, Dobson RCJ, Kennedy MA

Biochimie, 2020 Aug 14.
PMID: 32805304 DOI: 10.1016/j.biochi.2020.07.022

Next generation DNA sequencing and analysis of amplicons spanning the pharmacogene CYP2D6 suggested that the Nextera transposase used for fragmenting and providing sequencing priming sites displayed a targeting bias. This manifested as dramatically lower sequencing coverage at sites in the amplicon that appeared likely to form G-quadruplex structures. Since secondary DNA structures such as G-quadruplexes are abundant in the human genome, and are known to interact with many other proteins, we further investigated these sites of low coverage. Our investigation revealed that G-quadruplex structures are formed in vitro within the CYP2D6 pharmacogene at these sites, and G-quadruplexes can interact with the hyperactive Tn5 transposase (EZ-Tn5) with high affinity. These findings indicate that secondary DNA structures such as G-quadruplexes may represent preferential transposon integration sites and provide additional evidence for the role of G-quadruplex structures in transposition or viral integration processes.

Matched MeSH terms: High-Throughput Nucleotide Sequencing
Microbiota composition data for wild and captive bluestreak cleaner wrasse Labroides dimidiatus (Valenciennes, 1839)

Okomoda VT, Nurul ANA, Danish-Daniel AM, Oladimeji AS, Abol-Munafi AB, Alabi KI, et al.

Data Brief, 2020 Oct;32:106120.
PMID: 32817873 DOI: 10.1016/j.dib.2020.106120

The Labroides dimidiatus is known as the "doctor fish" because of its role in removing parasites and infectious pathogens from the body of other fishes. This important role played both in wild and captive conditions could represent a novel form of parasitic transmission process mediated by the cleaning activity of the fish. Yet, there is a paucity of data on the microflora associated with this fish which is important for tracking disease infection and generally monitoring the health status of the fish. This article, therefore, represents the first dataset for the microbiota composition of wild and captive L. dimidiatus. Wild fish samples and carriage water were gotten in Terengganu Malaysia around the corals of the Karah Island. The captive sample, however, was obtained from well-known ornamental fish suppliers in Terengganu Malaysia. Thereafter, bacteria present on the skin, in the stomach and the aquarium water were enumerated using culture-independent approaches and Next Generation Sequencing (NGS) technology. Data obtained from the three metagenomic libraries using NGS analysis gave 1,426,740 amplicon sequence reads which are composed of 508 operational taxonomic units (OTUs) for wild samples and 3,238,564 valid reads and 828 OTUs for captive samples. All sequence reads were deposited in the GeneBank (Accession numbers SAMN14260247, SAMN14260248, SAMN14260249, SAMN14260250, SAMN14260251, and SAMN14260252). The dataset presented is associated with the research article "16S rDNA-Based Metagenomic Analysis of Microbial Communities Associated with Wild Labroides dimidiatus From Karah Island, Terengganu, Malaysia" [1]. The microbiota data presented in this article can be used to monitor the health and wellbeing of the ornamental fish, especially under captivity, hence preventing possible cross-infection.

Matched MeSH terms: High-Throughput Nucleotide Sequencing

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