Displaying publications 61 - 80 of 216 in total

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  1. 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*
  2. Azaman SNA, Satharasinghe DA, Tan SW, Nagao N, Yusoff FM, Yeap SK
    Genes (Basel), 2020 09 25;11(10).
    PMID: 32992970 DOI: 10.3390/genes11101131
    Chlorella is a popular microalga with robust physiological and biochemical characteristics, which can be cultured under various conditions. The exploration of the small RNA content of Chlorella could improve strategies for the enhancement of metabolite production from this microalga. In this study, stress was introduced to the Chlorella sorokiniana culture to produce high-value metabolites such as carotenoids and phenolic content. The small RNA transcriptome of C. sorokiniana was sequenced, focusing on microRNA (miRNA) content. From the analysis, 98 miRNAs were identified in cultures subjected to normal and stress conditions. The functional analysis result showed that the miRNA targets found were most often involved in the biosynthesis of secondary metabolites, followed by protein metabolism, cell cycle, and porphyrin and chlorophyll metabolism. Furthermore, the biosynthesis of secondary metabolites such as carotenoids, terpenoids, and lipids was found mostly in stress conditions. These results may help to improve our understanding of regulatory mechanisms of miRNA in the biological and metabolic process of Chlorella species. It is important and timely to determine the true potential of this microalga species and to support the potential for genetic engineering of microalgae as they receive increasing focus for their development as an alternative source of biofuel, food, and health supplements.
    Matched MeSH terms: High-Throughput Nucleotide Sequencing/methods*
  3. 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
  4. Qiu B, Fang S, Ikhwanuddin M, Wong L, Ma H
    Mol Biol Rep, 2020 Apr;47(4):3011-3017.
    PMID: 32124169 DOI: 10.1007/s11033-020-05348-z
    In this study, we first conducted a genome survey assay for Sillago sihama by Illumina sequencing platform, and then developed 15 polymorphic microsatellite loci in a wild population. A total of 129.46 Gb raw data were obtained, of which 115.07 Gb were clean data, with a sequencing depth of 179.3-folds. This genome was estimated to be 522.6 Mb in size, with the heterozygosity, repeat content and GC content being 0.63%, 21% and 44%. A total of 630,028 microsatellites were identified from the genome, of which, dinucleotide repeat was the most abundant (56.80%), followed by mononucleotide repeat (30.23%). Furthermore, 60 pairs of primers were designed and synthesized based on microsatellite sequences, of which 15 were polymorphic in a wild population. A total of 91 alleles were found, with an average of 6.07 per locus. Number of alleles, observed and expected heterozygosity per locus ranged from two to 13, from 0.250 to 0.862, and from 0.396 to 0.901, respectively. Twelve loci were highly informative (PIC > 0.5), and the others were medium informative (0.25 
    Matched MeSH terms: High-Throughput Nucleotide Sequencing/methods
  5. Mohamed Yusoff A, Tan TK, Hari R, Koepfli KP, Wee WY, Antunes A, et al.
    Sci Rep, 2016 09 13;6:28199.
    PMID: 27618997 DOI: 10.1038/srep28199
    Pangolins are scale-covered mammals, containing eight endangered species. Maintaining pangolins in captivity is a significant challenge, in part because little is known about their genetics. Here we provide the first large-scale sequencing of the critically endangered Manis javanica transcriptomes from eight different organs using Illumina HiSeq technology, yielding ~75 Giga bases and 89,754 unigenes. We found some unigenes involved in the insect hormone biosynthesis pathway and also 747 lipids metabolism-related unigenes that may be insightful to understand the lipid metabolism system in pangolins. Comparative analysis between M. javanica and other mammals revealed many pangolin-specific genes significantly over-represented in stress-related processes, cell proliferation and external stimulus, probably reflecting the traits and adaptations of the analyzed pregnant female M. javanica. Our study provides an invaluable resource for future functional works that may be highly relevant for the conservation of pangolins.
    Matched MeSH terms: High-Throughput Nucleotide Sequencing/methods*
  6. Lee TY, Lai MI, Ramachandran V, Tan JA, Teh LK, Othman R, et al.
    Int J Lab Hematol, 2016 Aug;38(4):435-43.
    PMID: 27349818 DOI: 10.1111/ijlh.12520
    INTRODUCTION: Alpha thalassaemia is a highly prevalent disease globally and is a well-known public health problem in Malaysia. The deletional forms of the mutation are the most common forms found in alpha thalassaemia. The three most common deletional alpha thalassaemia found in this region include --(SEA) deletion, -α(3.7) rightward and -α(4.2) leftward deletions. The prevalence rate of triplication alpha cases such as ααα(anti3.7) and ααα(anti4.2) is not known in Malaysia although it plays a role in exacerbating the clinical phenotypes in beta thalassaemia carriers. Recently, there have been more reported cases of rare alpha thalassaemia mutations due to the advancement of molecular techniques involved in thalassaemia detections. Therefore, it is essential to develop a new method which allows the detection of different alpha thalassaemia mutations including the rare ones simultaneously and accurately.

    METHODS: The purpose of this study was to design an assay for the detection of triplications, common and rare deletional alpha thalassaemia using droplet digital PCR (ddPCR).

    RESULTS: This is a quantitative detection method to measure the changes of copy number which can detect deletions, duplications and triplications of the alpha globin gene simultaneously.

    CONCLUSION: In conclusion, ddPCR is an alternative method for rapid detection of alpha thalassaemia variants in Malaysia.

    Matched MeSH terms: High-Throughput Nucleotide Sequencing/methods*
  7. Ochiai E, Minaguchi K, Nambiar P, Kakimoto Y, Satoh F, Nakatome M, et al.
    Leg Med (Tokyo), 2016 Sep;22:58-61.
    PMID: 27591541 DOI: 10.1016/j.legalmed.2016.08.001
    The Y chromosomal haplogroup determined from single nucleotide polymorphism (SNP) combinations is a valuable genetic marker to study ancestral male lineage and ethical distribution. Next-generation sequencing has been developed for widely diverse genetics fields. For this study, we demonstrate 34 Y-SNP typing employing the Ion PGM™ system to perform haplogrouping. DNA libraries were constructed using the HID-Ion AmpliSeq™ Identity Panel. Emulsion PCR was performed, then DNA sequences were analyzed on the Ion 314 and 316 Chip Kit v2. Some difficulties became apparent during the analytic processes. No-call was reported at rs2032599 and M479 in six samples, in which the least coverage was observed at M479. A minor misreading occurred at rs2032631 and M479. A real time PCR experiment using other pairs of oligonucleotide primers showed that these events might result from the flanking sequence. Finally, Y haplogroup was determined completely for 81 unrelated males including Japanese (n=59) and Malay (n=22) subjects. The allelic divergence differed between the two populations. In comparison with the conventional Sanger method, next-generation sequencing provides a comprehensive SNP analysis with convenient procedures, but further system improvement is necessary.
    Matched MeSH terms: High-Throughput Nucleotide Sequencing/methods
  8. 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*
  9. Soliman AM, Das S, Teoh SL
    Int J Mol Sci, 2021 Jul 13;22(14).
    PMID: 34299097 DOI: 10.3390/ijms22147470
    Multiple myeloma (MM) is considered to be the second most common blood malignancy and it is characterized by abnormal proliferation and an accumulation of malignant plasma cells in the bone marrow. Although the currently utilized markers in the diagnosis and assessment of MM are showing promising results, the incidence and mortality rate of the disease are still high. Therefore, exploring and developing better diagnostic or prognostic biomarkers have drawn global interest. In the present review, we highlight some of the recently reported and investigated novel biomarkers that have great potentials as diagnostic and/or prognostic tools in MM. These biomarkers include angiogenic markers, miRNAs as well as proteomic and immunological biomarkers. Moreover, we present some of the advanced methodologies that could be utilized in the early and competent diagnosis of MM. The present review also focuses on understanding the molecular concepts and pathways involved in these biomarkers in order to validate and efficiently utilize them. The present review may also help in identifying areas of improvement for better diagnosis and superior outcomes of MM.
    Matched MeSH terms: High-Throughput Nucleotide Sequencing/methods*
  10. 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
  11. Gormus U, Chaubey A, Shenoy S, Wong YW, Chan LY, Choo BP, et al.
    Curr Issues Mol Biol, 2021 Aug 17;43(2):958-964.
    PMID: 34449543 DOI: 10.3390/cimb43020068
    Background: Rolling-circle replication (RCR) is a novel technology that has not been applied to cell-free DNA (cfDNA) testing until recently. Given the cost and simplicity advantages of this technology compared to other platforms currently used in cfDNA analysis, an assessment of RCR in clinical laboratories was performed. Here, we present the first validation study from clinical laboratories utilizing RCR technology. Methods: 831 samples from spontaneously pregnant women carrying a singleton fetus, and 25 synthetic samples, were analyzed for the fetal risk of trisomy 21 (T21), trisomy 18 (T18) and trisomy 13 (T13), by three laboratories on three continents. All the screen-positive pregnancies were provided post-test genetic counseling and confirmatory diagnostic invasive testing (e.g., amniocentesis). The screen-negative pregnancies were routinely evaluated at birth for fetal aneuploidies, using newborn examinations, and any suspected aneuploidies would have been offered diagnostic testing or confirmed with karyotyping. Results: The study found rolling-circle replication to be a highly viable technology for the clinical assessment of fetal aneuploidies, with 100% sensitivity for T21 (95% CI: 82.35-100.00%); 100.00% sensitivity for T18 (71.51-100.00%); and 100.00% sensitivity for T13 analyses (66.37-100.00%). The specificities were >99% for each trisomy (99.7% (99.01-99.97%) for T21; 99.5% (98.62-99.85%) for T18; 99.7% (99.03-99.97%) for T13), along with a first-pass no-call rate of 0.93%. Conclusions: The study showed that using a rolling-circle replication-based cfDNA system for the evaluation of the common aneuploidies would provide greater accuracy and clinical utility compared to conventional biochemical screening, and it would provide comparable results to other reported cfDNA methodologies.
    Matched MeSH terms: High-Throughput Nucleotide Sequencing/methods*
  12. Higashino A, Sakate R, Kameoka Y, Takahashi I, Hirata M, Tanuma R, et al.
    Genome Biol, 2012;13(7):R58.
    PMID: 22747675 DOI: 10.1186/gb-2012-13-7-r58
    The genetic background of the cynomolgus macaque (Macaca fascicularis) is made complex by the high genetic diversity, population structure, and gene introgression from the closely related rhesus macaque (Macaca mulatta). Herein we report the whole-genome sequence of a Malaysian cynomolgus macaque male with more than 40-fold coverage, which was determined using a resequencing method based on the Indian rhesus macaque genome.
    Matched MeSH terms: High-Throughput Nucleotide Sequencing/methods*
  13. Wong LP, Ong RT, Poh WT, Liu X, Chen P, Li R, et al.
    Am J Hum Genet, 2013 Jan 10;92(1):52-66.
    PMID: 23290073 DOI: 10.1016/j.ajhg.2012.12.005
    Whole-genome sequencing across multiple samples in a population provides an unprecedented opportunity for comprehensively characterizing the polymorphic variants in the population. Although the 1000 Genomes Project (1KGP) has offered brief insights into the value of population-level sequencing, the low coverage has compromised the ability to confidently detect rare and low-frequency variants. In addition, the composition of populations in the 1KGP is not complete, despite the fact that the study design has been extended to more than 2,500 samples from more than 20 population groups. The Malays are one of the Austronesian groups predominantly present in Southeast Asia and Oceania, and the Singapore Sequencing Malay Project (SSMP) aims to perform deep whole-genome sequencing of 100 healthy Malays. By sequencing at a minimum of 30× coverage, we have illustrated the higher sensitivity at detecting low-frequency and rare variants and the ability to investigate the presence of hotspots of functional mutations. Compared to the low-pass sequencing in the 1KGP, the deeper coverage allows more functional variants to be identified for each person. A comparison of the fidelity of genotype imputation of Malays indicated that a population-specific reference panel, such as the SSMP, outperforms a cosmopolitan panel with larger number of individuals for common SNPs. For lower-frequency (<5%) markers, a larger number of individuals might have to be whole-genome sequenced so that the accuracy currently afforded by the 1KGP can be achieved. The SSMP data are expected to be the benchmark for evaluating the value of deep population-level sequencing versus low-pass sequencing, especially in populations that are poorly represented in population-genetics studies.
    Matched MeSH terms: High-Throughput Nucleotide Sequencing*
  14. Ong WD, Voo LY, Kumar VS
    PLoS One, 2012;7(10):e46937.
    PMID: 23091603 DOI: 10.1371/journal.pone.0046937
    BACKGROUND: Pineapple (Ananas comosus var. comosus), is an important tropical non-climacteric fruit with high commercial potential. Understanding the mechanism and processes underlying fruit ripening would enable scientists to enhance the improvement of quality traits such as, flavor, texture, appearance and fruit sweetness. Although, the pineapple is an important fruit, there is insufficient transcriptomic or genomic information that is available in public databases. Application of high throughput transcriptome sequencing to profile the pineapple fruit transcripts is therefore needed.

    METHODOLOGY/PRINCIPAL FINDINGS: To facilitate this, we have performed transcriptome sequencing of ripe yellow pineapple fruit flesh using Illumina technology. About 4.7 millions Illumina paired-end reads were generated and assembled using the Velvet de novo assembler. The assembly produced 28,728 unique transcripts with a mean length of approximately 200 bp. Sequence similarity search against non-redundant NCBI database identified a total of 16,932 unique transcripts (58.93%) with significant hits. Out of these, 15,507 unique transcripts were assigned to gene ontology terms. Functional annotation against Kyoto Encyclopedia of Genes and Genomes pathway database identified 13,598 unique transcripts (47.33%) which were mapped to 126 pathways. The assembly revealed many transcripts that were previously unknown.

    CONCLUSIONS: The unique transcripts derived from this work have rapidly increased of the number of the pineapple fruit mRNA transcripts as it is now available in public databases. This information can be further utilized in gene expression, genomics and other functional genomics studies in pineapple.

    Matched MeSH terms: High-Throughput Nucleotide Sequencing*
  15. Loh KH, Shao KT, Chen HM, Chen CH, Chong VC, Loo PL, et al.
    Mitochondrial DNA A DNA Mapp Seq Anal, 2016 11;27(6):4230-4231.
    PMID: 26000942
    In this study, the complete mitogenome sequence of the Zebra moray, Gymnomuraena zebra (Anguilliformes: Muraenidae) has been sequenced by the next-generation sequencing method. The assembled mitogenome consisting of 16,576 bp includes 13 protein coding genes, 22 transfer RNAs, and two ribosomal RNAs genes. The overall base composition of Zebra moray is 30.2% for A, 26.8% for C, 17.2% for G, and 25.8% for T and show 80% identities to Kidako moray, Gymnothorax kidako. The complete mitogenome of the Zebra moray provides an essential and important DNA molecular data for further phylogeography and evolutionary analysis for moray eel phylogeny.
    Matched MeSH terms: High-Throughput Nucleotide Sequencing/methods
  16. Loh KH, Shao KT, Chen HM, Chen CH, Loo PL, Hui AT, et al.
    PMID: 26016872 DOI: 10.3109/19401736.2015.1030629
    In this study, the complete mitogenome sequence of the longfang moray, Enchelynassa canina (Anguilliformes: Muraenidae) has been sequenced by the next-generation sequencing method. The length of the assembled mitogenome is 16,592 bp, which includes 13 protein coding genes, 22 transfer RNAs, and 2 ribosomal RNAs genes. The overall base composition of longfang moray is 28.4% for A, 28.0% for C, 18.4% for G, 25.1% for T, and show 82% identities to Kidako moray, Gymnothorax kidako. The complete mitogenome of the longfang moray provides an essential and important DNA molecular data for further phylogeography and evolutionary analysis for moray eel phylogeny.
    Matched MeSH terms: High-Throughput Nucleotide Sequencing*
  17. Raabe CA, Tang TH, Brosius J, Rozhdestvensky TS
    Nucleic Acids Res, 2014 Feb;42(3):1414-26.
    PMID: 24198247 DOI: 10.1093/nar/gkt1021
    High-throughput RNA sequencing (RNA-seq) is considered a powerful tool for novel gene discovery and fine-tuned transcriptional profiling. The digital nature of RNA-seq is also believed to simplify meta-analysis and to reduce background noise associated with hybridization-based approaches. The development of multiplex sequencing enables efficient and economic parallel analysis of gene expression. In addition, RNA-seq is of particular value when low RNA expression or modest changes between samples are monitored. However, recent data uncovered severe bias in the sequencing of small non-protein coding RNA (small RNA-seq or sRNA-seq), such that the expression levels of some RNAs appeared to be artificially enhanced and others diminished or even undetectable. The use of different adapters and barcodes during ligation as well as complex RNA structures and modifications drastically influence cDNA synthesis efficacies and exemplify sources of bias in deep sequencing. In addition, variable specific RNA G/C-content is associated with unequal polymerase chain reaction amplification efficiencies. Given the central importance of RNA-seq to molecular biology and personalized medicine, we review recent findings that challenge small non-protein coding RNA-seq data and suggest approaches and precautions to overcome or minimize bias.
    Matched MeSH terms: High-Throughput Nucleotide Sequencing/methods*
  18. Lim CC, Lim TS
    Int J Biol Macromol, 2025 Feb;290:140037.
    PMID: 39828167 DOI: 10.1016/j.ijbiomac.2025.140037
    Lymphatic filariasis is caused by infections of thread-like filarial worms, namely Wuchereria bancrofti, Brugia Malayi and Brugia timori. However, in-depth analysis of the antibody repertoire against Lymphatic filariasis is lacking. Using high-throughput sequencing of antibody repertoires, immunome analysis of IgG (LG) and IgM (LM) repertoires were studied. Despite significant differences between LG and LM in V(D)J gene usage, IGHV4-34, IGHV6-1, IGHD3-10 and IGHJ4 were preferred in both repertoires. The CDR3 in the LG repertoire showed a longer length than LM. Higher SHM level were observed in LG sequences and presence of oligoclonal sequences indicates the extent of clonal expansion. The prevalence of rare clonotypes in LM repertoire depicts the high clonal diversity when compared to LG repertoire. Monoclonal antibodies against closely related parasitic infections were present within the LG repertoire suggesting that immune repertoires may not be as exclusive and biased against the target infection as initially thought. The characterization of the immune repertoire can provide critical insight into the antibody response patterns in disease state, antibody generation process during infections and future antibody designs.
    Matched MeSH terms: High-Throughput Nucleotide Sequencing*
  19. Md-Zain BM, Wan-Mustafa WAS, Tingga RCT, Gani M, Mohd-Ridwan AR
    J Med Primatol, 2025 Apr;54(2):e70009.
    PMID: 40012216 DOI: 10.1111/jmp.70009
    BACKGROUND: The gut microbiota plays a vital role in primates' overall health and well-being, including small apes (Hylobatidae). The symbiotic relationships between bacteria and the gut aid food digestion, maintain host health, and help them adapt to their environment, including captive conditions. Despite being listed as endangered in the International Union for Conservation of Nature (IUCN) red list category, molecular studies on the small ape's gut microbiome are limited compared to other primates. This study aimed to characterize the gut microbiota of captive small apes at Zoo Taiping and Night Safari, Peninsular Malaysia, by evaluating their microbial communities.

    METHODS: Seven fecal samples from Hylobatidae (white-handed gibbon and siamang) were collected, and the bacteria therein were successfully isolated and subjected to high-throughput sequencing of the 16S rRNA gene.

    RESULTS: The acquired amplicon sequence variants (ASVs) were successfully classified into 17 phyla, 82 families, 164 genera, and 43 species of microbes. Each small ape exhibited a unique gut microbiota profile. The phyla Bacteroidota and Firmicutes were dominant in each individual. Environmental conditions and host genetics are among the factors that influence the small ape's gut microbiome composition.

    CONCLUSIONS: These findings provide valuable insights into the gut microbiota composition of small apes at Zoo Taiping and Night Safari, thus contributing to the health management and welfare efforts of small apes in captivity.

    Matched MeSH terms: High-Throughput Nucleotide Sequencing/veterinary
  20. Chin KCJ, Taylor TD, Hebrard M, Anbalagan K, Dashti MG, Phua KK
    BMC Genomics, 2017 Oct 31;18(1):836.
    PMID: 29089020 DOI: 10.1186/s12864-017-4212-6
    BACKGROUND: Typhoid fever is an acute systemic infection of humans caused by Salmonella enterica subspecies enterica serovar Typhi (S. Typhi). In chronic carriers, the bacteria survive the harsh environment of the gallbladder by producing biofilm. The phenotype of S. Typhi biofilm cells is significantly different from the free-swimming planktonic cells, and studies have shown that they are associated with antibiotic resistance, immune system evasion, and bacterial persistence. However, the mechanism of this transition and the events leading to biofilm formation are unknown. High throughput sequencing was performed to identify the genes involved in biofilm formation and to postulate the mechanism of action.

    RESULTS: Planktonic S. Typhi cells were cultured using standard nutrient broth whereas biofilm cells were cultured in a stressful environment using high shearing-force and bile to mimic the gallbladder. Sequencing libraries were prepared from S. Typhi planktonic cells and mature biofilm cells using the Illumina HiSeq 2500 platform, and the transcriptome data obtained were processed using Cufflinks bioinformatics suite of programs to investigate differential gene expression between the two phenotypes. A total of 35 up-regulated and 29 down-regulated genes were identified. The identities of the differentially expressed genes were confirmed using NCBI BLAST and their functions were analyzed. The results showed that the genes associated with metabolic processes and biofilm regulations were down-regulated while those associated with the membrane matrix and antibiotic resistance were highly up-regulated.

    CONCLUSIONS: It is proposed that the biofilm phenotype of S. Typhi allows the bacteria to increase production of the membrane matrix in order to serve as a physical shield and to adhere to surfaces, and enter an energy conservation state in response to the stressful environment. Conversely, the planktonic phenotype allows the bacteria to produce flagella and increase metabolic activity to enable the bacteria to migrate and form new colonies of infection. This data provide a basis for further studies to uncover the mechanism of biofilm formation in S. Typhi and to discover novel genes or pathways associated with the development of the typhoid carrier state.

    Matched MeSH terms: High-Throughput Nucleotide Sequencing
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