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At least 19 recordsLinked to original sources

PubMed2026

New insights on Plasmodium gene expression from direct RNA sequencing.

Oxford Nanopore Technology (ONT) direct RNA sequencing enables the sequencing of native RNA molecules without cDNA conversion. The long-read approach captures full-length reads spanning entire genes and has transformed the study of gene expression in Plasmodium parasites by enabling analysis of untranslated regions, isoforms, and alternative splicing. In addition, ONT provides unique insights into non-coding RNAs, RNA modifications, and polyadenylated tail dynamics, which are expanding our understanding of post-transcriptional regulation in Plasmodium, including processes beyond translational repression in gametocytes and sporozoites. Here, we discuss the past and future applications of direct RNA sequencing in Plasmodium research and highlight its advantages, limitations, and future prospects.

Oxford Nanopore Technology↗

PubMed2026

ModiCal: A Targeted Calibration Workflow for Site-Specific m5C Validation by Nanopore Direct RNA Sequencing.

Accurate identification of RNA 5-methylcytidine (m5C) at the single-nucleotide resolution remains a central challenge in nanopore direct RNA sequencing (DRS). Current global scanning and modification-aware basecalling methods enable transcriptome-wide profiling but often yield high false-positive rates and lack site-specific accuracy. To address this, we repurposed ModiDeC, originally a de novo multimodification classifier, into a targeted, high-precision validation tool for RNA modification sites with prior biochemical knowledge. This was implemented through a three-step calibration workflow that alternates between biochemical and computational modules using the well-characterized m5C2278 site in 25S rRNA as a starting point. Baseline training uses short synthetic RNAs carrying either a methylated or unmodified C2278 as ground truth, followed by IVT-derived calibration and validation in methyltransferase knockout yeast. The baseline model accurately detected the bona fide m5C2278 site but initially produced off-target predictions. Iterative retraining with unmodified IVT signals progressively reduced and ultimately eliminated false positives while maintaining a strong signal at the bona fide site. The final model retained enzyme-dependent detection in wild-type versus knockout yeast and, when explicitly targeted, was also able to detect the second rRNA site, C2870, which remained invisible in the initial analysis. Application to native human prerRNA processing intermediates further resolved two distinct m5C deposition regimes on 28S rRNA, while generalization to dengue virus genomic RNA confirmed that the same calibration logic transfers across diverse RNA contexts. Together, this study establishes a reproducible and transferable framework that integrates biochemical validation with iterative neural network refinement, providing a route toward reliable site-specific m5C confirmation by nanopore direct RNA sequencing.

RNA Methylation↗

PubMed2026

DirectASRM: uncovering allele-specific post-transcriptional RNA modifications through direct RNA sequencing.

SUMMARY: We developed DirectASRM, a comprehensive database for the systematic identification, integration, and annotation of allele-specific RNA modifications (ASRMs) from direct RNA sequencing data. DirectASRM enables single-base, transcript-level detection of ASRMs across multiple RNA modification types, diverse organisms and condition-specific contexts. The database further evaluates the confidence of each ASRM-SNP pair association within isoform context by jointly considering statistical evidence of allelic modification imbalance and independent support from external next-generation sequencing (NGS) - based RNA modification resources. DirectASRM also provides extensive functional annotations for ASRMs and their associated variants, including intra-sample transcript-level allele-specific expression (ASE) and allele-specific splicing, as well as additional post-transcriptional regulatory features such as miRNA binding, circRNA, RNA-protein interactions, and disease relevance. Overall, DirectASRM serves as a comprehensive resource that supports systematic investigation of the potential functional impact of genetic variants in epitranscriptomic regulation. AVAILABILITY AND IMPLEMENTATION: DirectASRM database is freely accessible at http://modinfor.com/DirectASRM/. DirectASRM pipeline is available at GitHub (https://github.com/jiayin1101/DirectASRM_pipeline) and Zenodo (DOI: https://doi.org/10.5281/zenodo.19876077).

Alleles↗

PubMed2026

Characterization of METTL3/14-mediated m6A modification in human transcriptome using Nanopore direct RNA sequencing.

Post-transcriptional RNA modifications modulate diverse aspects of RNA metabolism. N6-methyladenosine (m6A), one of the most abundant internal RNA modifications, is deposited by the core methyltransferase complex, METTL3 and METTL14. Oxford Nanopore Technologies (ONT) platform permits direct, single RNA molecule sequencing while preserving native modifications. However, without rigorous benchmarking, the accuracy and reproducibility of modification detection remain uncertain. Here, we leveraged ONT to comprehensively profile bona fide m6A modifications in cellular RNAs at single-nucleotide resolution by integrating two direct RNA sequencing chemistries (RNA002 and RNA004) with the m6Anet and Dorado modification-detection models. We independently depleted METTL3 and METTL14 in human cells and rigorously validated modification calls through several assays and independent orthogonal methods (GLORI and miCLIP). We find that Dorado detected a higher number of m6A events and enabled simultaneous detection of other RNA modifications (5-methylcytosine, pseudouridine, and inosine). Pairing Dorado with an in vitro transcribed, unmodified control under stringent filtering, we provide compelling evidence supporting a global reduction in m6A sites and stoichiometry within coding sequences and across genes, particularly in highly modified genes and sites, and at consensus DRACH motifs. We report a differential and complex regulation of modified transcripts, accompanied by a global reduction in poly(A) tail length. Notably, METTL3 and METTL14 depletion produced distinct transcript-specific effects, supporting non-redundant roles within the m6A writer complex. Together, our study illustrates a notable advancement of ONT capabilities and establishes a robust transcriptome-wide framework for RNA modification detection, thereby laying the groundwork for exploring the contribution of METTL3/METTL14 to cellular functions and disease.

Humans↗

PubMed2019

Direct RNA nanopore sequencing of full-length coronavirus genomes provides novel insights into structural variants and enables modification analysis.

Sequence analyses of RNA virus genomes remain challenging owing to the exceptional genetic plasticity of these viruses. Because of high mutation and recombination rates, genome replication by viral RNA-dependent RNA polymerases leads to populations of closely related viruses, so-called "quasispecies." Standard (short-read) sequencing technologies are ill-suited to reconstruct large numbers of full-length haplotypes of (1) RNA virus genomes and (2) subgenome-length (sg) RNAs composed of noncontiguous genome regions. Here, we used a full-length, direct RNA sequencing (DRS) approach based on nanopores to characterize viral RNAs produced in cells infected with a human coronavirus. By using DRS, we were able to map the longest (∼26-kb) contiguous read to the viral reference genome. By combining Illumina and Oxford Nanopore sequencing, we reconstructed a highly accurate consensus sequence of the human coronavirus (HCoV)-229E genome (27.3 kb). Furthermore, by using long reads that did not require an assembly step, we were able to identify, in infected cells, diverse and novel HCoV-229E sg RNAs that remain to be characterized. Also, the DRS approach, which circumvents reverse transcription and amplification of RNA, allowed us to detect methylation sites in viral RNAs. Our work paves the way for haplotype-based analyses of viral quasispecies by showing the feasibility of intra-sample haplotype separation. Even though several technical challenges remain to be addressed to exploit the potential of the nanopore technology fully, our work illustrates that DRS may significantly advance genomic studies of complex virus populations, including predictions on long-range interactions in individual full-length viral RNA haplotypes.

Cell Line↗

PubMed2024

Nanopore sequencing to detect A-to-I editing sites.

Adenosine-to-inosine (A-to-I) RNA editing, mediated by the ADAR family of enzymes, is pervasive in metazoans and functions as an important mechanism to diversify the proteome and control gene expression. Over the years, there have been multiple efforts to comprehensively map the editing landscape in different organisms and in different disease states. As inosine (I) is recognized largely as guanosine (G) by cellular machineries including the reverse transcriptase, editing sites can be detected as A-to-G changes during sequencing of complementary DNA (cDNA). However, such an approach is indirect and can be confounded by genomic single nucleotide polymorphisms (SNPs) and DNA mutations. Moreover, past studies rely primarily on the Illumina platform, which generates short sequencing reads that can be challenging to map. Recently, nanopore direct RNA sequencing has emerged as a powerful technology to address the issues. Here, we describe the use of the technology together with deep learning models that we have developed, named Dinopore (Detection of inosine with nanopore sequencing), to interrogate the A-to-I editome of any organism.

Inosine↗

PubMed2025

Degradation of ribosomal RNA during Plasmodium falciparum gametocytogenesis.

The life cycle of Plasmodium falciparum is characterized by complex regulatory changes that allow adaptation of the parasites to different environmental conditions, which are especially pronounced during transmission between the mammalian host and the insect vector. Previous studies have shown that P. falciparum uses three types of ribosomal RNAs (rRNA A, S1, and S2 types) at different stages of its life cycle. We used Oxford Nanopore Technologies direct RNA sequencing to investigate the dynamics of rRNA usage throughout the parasite's intraerythrocytic development, as well as in salivary gland sporozoites. Our study revealed a preponderance of A-type rRNAs during the intraerythrocytic cycle and gametocytogenesis, while S-type rRNAs slowly increase in abundance in mosquito stages starting 3 days post-infection. Salivary gland sporozoites showed an even proportion of all rRNA types. By examining the length distributions of rRNA molecules, we detected an extensive and specific degradation of rRNAs during gametocytogenesis, starting in stage II gametocytes and continuing until the final stages of gametocyte development. We hypothesize that rRNA degradation may be linked to the global translational repression and metabolic quiescence described in stage V gametocytes, similar to mechanisms observed in bacterial and eukaryotic stress responses.IMPORTANCEOur study uses Oxford Nanopore direct RNA sequencing of tightly synchronized blood-stage Plasmodium falciparum parasites to investigate the expression of ribosomal RNAs during asexual and sexual development. P. falciparum utilizes distinct types of rRNA during its development. However, due to the challenges of differentiating these highly similar molecules, their regulation and the mechanism underlying the switch between rRNA types remain unclear. We observe significant rRNA degradation in mature gametocytes, leading us to propose that this potentially leads to a reduced number of functional ribosomes when parasites become quiescent and translation is repressed.

Plasmodium falciparum↗

PubMed2026

Long-read transcriptomics corrects Trichomonas vaginalis intron annotations and refines transcript-end features.

BACKGROUND: Trichomonas vaginalis causes the most prevalent non-viral sexually transmitted infection worldwide. Despite its large genome (181.5 Mb; 36,310 predicted protein-coding genes in NYU_TvagG3_2), intron annotations remain limited and inconsistently validated. A recent short-read RNA-seq study reported 63 putative active introns, but short reads can misassign splice boundaries and cannot resolve complete transcript structures. METHODS: We integrated Oxford Nanopore direct RNA sequencing (DRS), ONT cDNA long-read sequencing, and Illumina RNA-seq to refine intron annotations, transcript-end features, and UTR boundaries in T. vaginalis. Candidate introns were validated by targeted PCR and Sanger sequencing, and representative splicing events were further assessed using public SRA datasets. RESULTS: Starting from 31 historically annotated introns, motif-guided long-read screening and orthogonal validation identified 17 additional validated introns, increasing the curated set to 48 confirmed introns. Among these 17 events, three were previously unrecognized in the current NYU_TvagG3_2 reference annotation. We also corrected five reported loci, including two false-positive introns, two splice-coordinate misannotations, and one gene-sequence error. DRS further supported transcript termination site mapping, UAAA polyadenylation-signal profiling relative to poly(A) addition sites, and single-molecule poly(A)-tail estimation. StringTie mixed-mode assemblies provided updated UTR boundaries for intron-bearing transcripts and transcripts without curated introns. CONCLUSIONS: This study provides a rigorously validated, long-read-refined resource of intron annotations, UTR boundaries, and UAAA-guided transcript-end features for T. vaginalis, together with a reproducible workflow for non-model protists. These refinements improve the current reference annotation and support future studies of functional genomics, parasite biology, pathogenesis, and diagnostic development.

Trichomonas vaginalis↗

PubMed1979

Direct chemical method for sequencing RNA.

Four different base-specific chemical reactions generate a means of directly sequencing RNA terminally labeled with 32P. After a partial, specific modification of each kind of RNA base, an amine-catalyzed strand scission generates labeled fragments whose lengths determine the position of each nucleotide in the sequence. Dimethyl sulfate modifies guanosine. Diethyl pyrocarbonate attacks primarily adenosine. Hydrazine attacks uridine and cytidine, but salt suppresses the reaction with uridine. In all cases, aniline induces a subsequent strand scission. The electrophoretic fractionation of the labeled fragments on a polyacrylamide gel, followed by autoradiography, determines the RNA sequence. RNA labeled at the 3' end yields clean cleavage patterns for each purine and pyrimidine and allows a determination of the entire RNA sequence out to 100-200 bases from the labeled terminus.

Base Sequence↗

PubMed1978

Complete nucleotide sequence of a 16S ribosomal RNA gene from Escherichia coli.

The complete nucleotide sequence of the 16S RNA gene from the rrnB cistron of Escherichia coli has been determined by using three rapid DNA sequencing methods. Nearly all of the structure has been confirmed by two to six independent sequence determinations on both DNA strands. The length of the 16S rRNA chain inferred from the DNA sequence is 1541 nucleotides, in close agreement with previous estimates. We note discrepancies between this sequence and the most recent version of it reported from direct RNA sequencing [Ehresmann, C., Stiegler, P., Carbon, P. & Ebel, J.P. (1977) FEBS Lett. 84, 337-341]. A few of these may be explained by heterogeneity among 16S rRNA sequences from different cistrons. No nucleotide sequences were found in the 16S rRNA gene that cannot be reconciled with RNase digestion products of mature 16S rRNA.

Base Sequence↗

PubMed2026

A critical appraisal of base-resolution m6A profiling techniques.

N6-methyladenosine (m6A) is the most prevalent internal modification in eukaryotic mRNA, influencing RNA fate and gene regulation. Early antibody-based approaches enabled transcriptome-wide profiling but lacked resolution and quantitative accuracy. Newer approaches now achieve base-resolution m6A detection using improved crosslinking, chemical or enzymatic conversion, and single-molecule sequencing. Antibody-free methods provide quantitative stoichiometry from minimal input, while nanopore direct RNA sequencing offers real-time, single-molecule readouts across entire transcriptomes. Collectively, these methods form a versatile toolkit that integrates global mapping with precise site-level analysis, advancing knowledge of context-dependent m6A regulation in physiology and disease. This review compares their principles, strengths, and limitations to guide method selection and highlight how next-generation epitranscriptomic tools are paving the way for clinical and therapeutic applications.

Humans↗

PubMed1978

Coincidence of the promoter and capped 5' terminus of RNA from the adenovirus 2 major late transcription unit.

During the late stage of adenovirus 2 infection, RNA chains are initiated at a site near coordinate 16 (Evans et al., 1977) and transcribed approximately 30,000 nucleotides to the far end of the genome at coordinate 100. Late mRNAs processed from these transcripts contain a common spliced tripartite leader (Berget, Moore and Sharp, 1977; Chow et al., 1977a) encoded at approximately 16, 20 and 27, and protein coding sequences which map downstream. This report maps the late promoter and the capped 5' end of nuclear and cytoplasmic RNAs from this transciption unit, and analyzes their structures. We show that nascent RNA chains pulse-labeled in vivo are initiated at coordinate 16.5 +/- 0.5 and contain the sequences intervening between the leader segments. We map the capped 5' terminus of late nuclear transcripts at a site between 16.4 and 16.6 by aligning T1 RNAase oligonucleotides from nuclear RNA with the DNA sequence of the promoter region. The structure of the first eleven residues of the capped 5' terminus of late mRNA was determined by direct RNA sequencing. This structure corresponds exactly to a DNA sequence at coordinate 16.4 and precisely positions the mRNA cap template within the promoter region. These results suggest that the promoter and the cap template sites are coincident, and that the initiating residues of the primary transcript are precursors of the capped 5' end of mRNA. Residues removed from transcripts by splicing were identified. These plus caps were detected in large polyadenylated nuclear RNA, indicating that capping and polyadenylation can occur on unspliced molecules. Residues retained in the mRNA first leader contain a nine residue sequence adjacent to the cap which is complementary to the 3' end of 18S rRNA, suggesting that the first leader functions in ribosome binding. Nucleotide sequences from the promoter region are compared with cellular counterparts. Strong homologies at cap sites and splice points suggest that for the noted cases, the virus and cell share closely related mechanisms for mRNA 5' end synthesis and splicing.

Adenoviruses, Human↗

PubMed2026

Nanopore Sequencing Reveals rRNA Modification Changes in Human Cells Experiencing Oxidative or Inflammatory Stress.

Ribosomal RNA (rRNA) modifications are tuned to regulate protein synthesis; however, their temporal dynamics during oxidative or inflammatory stress remain poorly understood. Nanopore direct RNA sequencing using Dorado v5.2.0 modification-aware models for the data analysis was employed to map human rRNA epitranscriptomic marks in a cell line undergoing oxidative stress, inflammatory stress, or ferroptosis. Oxidative stress triggered a global trend of decreased modification occupancy in which six modifications shifted significantly over 48 h, particularly, 18S Ψ573 and 18S m6A1832. Conversely, inflammatory stress induced a complex response involving an acute pulse of hypermodifications at 28S Um1773 and 28S Ψ1779, for example, and chronic hypomodification at specific target sites (e.g., 18S Gm1328 and 28S Gm4228). In this work, the pseudouridine modifications 28S Ψ4296 and 28S Ψ4353 were identified as "universal stress markers" that decreased under all stressors studied, including ferroptosis. Mapping these changes onto the ribosome structure revealed that they reside in functional regions such as the decoding center and A-site finger, supporting a role in functional ribosome reprogramming during stress. Analysis of mitochondrial rRNA (mt-rRNA) revealed modification shifts within the peptidyl transferase center, suggesting a mechanism to attenuate mitochondrial translation during chronic stress. This work demonstrates that oxidative and inflammatory stress drive distinct, time-resolved remodeling of the human rRNA epitranscriptome and provides a framework for using rRNA modifications as biomarkers of cellular health during oxidative or inflammatory stress exposure.

Humans↗

PubMed2026

Functional chimeric mRNAs encode proteins in mammalian immunity.

Individual mammalian mRNAs and proteins are typically believed to originate from single genomic loci, with isoform diversity arising through cis-splicing of pre-mRNA. Whether mRNA from distant genes can undergo trans-splicing to generate functionally relevant chimeric transcripts has remained unclear. Here we develop a pipeline combining long-read direct RNA sequencing with non-targeted and targeted validation to identify chimeric transcripts in macrophages. Chromatin conformation capture studies reveal that inflammation induces interchromosomal DNA interactions, positioning parent genes proximally to facilitate the formation of chimeric mRNA. Notably, we identify a protein-coding chimeric mRNA representing a fusion between the pore-forming protein gasdermin D (GSDMD)1,2 and a C-terminal domain translated out of frame from Tmem106a (Gsdmd-Tmem106a) in mice. We show that inflammasome priming upregulates Gsdmd-Tmem106a, with the protein localizing to the plasma membrane. After activation of the inflammasome, GSDMD-TMEM106A directly interacts with canonical GSDMD N termini to accelerate and enhance pore formation and IL-1β release. Finally, we show that GSDMD-TMEM106A balances host defence and immunopathology in vivo: its loss protects against lethal sepsis but compromises antibacterial defence, whereas overexpression enhances host protection while increasing sepsis lethality. We establish that protein-coding chimeric mRNAs formed by regulated transcript fusion events are operative during inflammation and immunity.

Journal Article↗

PubMed2026

Maternal immune activation perturbs the brain epitranscriptome.

Maternal immune activation (MIA) results in abnormal fetal neurodevelopment and an increased risk of neurodevelopmental disorders. Altered RNA translation has been implicated in the pathophysiology of MIA-associated neurodevelopmental deficits, but more precise mechanisms underlying disruption in RNA metabolism are lacking. Here, we characterize key components of the RNA epitranscriptomic machinery, which refers to the set of reversible chemical modifications on RNA molecules that influence RNA function, including translation, stability, splicing, and localization. Using spatial transcriptomics, we define cell type- and brain region-specific distribution of epitranscriptome regulators in the developing mouse brain. We also use direct RNA sequencing to define how MIA changes the brain epitranscriptome landscape. We identify the demethylase FTO as being notably perturbed in the context of MIA. Using pharmacological and genetic approaches, we target FTO to ameliorate behavioral phenotypes in MIA offspring. In total, this work expands upon mechanisms of translational misregulation in MIA and identifies new targets for therapeutic manipulation.

Animals↗

PubMed1979

Globin synthesis and erythroid differentiation in a Friend cell variant deficient in heme synthesis.

Friend erythroleukemia cells of line Fw are noninducible for hemoglobin synthesis by dimethyl sulfoxide, butyric acid, and other agents. However, these agents were found to induce hemoglobin synthesis if the cells were also treated with exogenous hemin. Butyric acid (or, to a lesser extent, dimethyl sulfoxide) by itself induced accumulation of the erythroid-specific membrane protein spectrin. The basis of the control of globin gene expression by hemin was investigated. Hemin does not control globin and total protein synthesis via the action of the hemin-controlled repressor. Rather, hemin, alone or in combination with dimethyl sulfoxide, induces accumulation of globin mRNA and nuclear globin RNA sequences. Direct measurements of heme synthesis indicate that Fw cells may be significantly deficient in heme metabolism.

Animals↗

PubMed2025

The UTRs of Leishmania donovani vary in length and are enriched in potential regulatory structures.

Leishmania spp. regulate gene expression largely post-transcriptionally, yet untranslated regions (UTRs) remain poorly delineated. We generated high-quality genome and transcriptome datasets for Leishmania donovani strain 1S2D (Ld1S) by combining PacBio HiFi de novo assembly with Oxford Nanopore direct RNA sequencing of promastigotes and axenic amastigotes. The genome assembly consists of 65 scaffolds totaling ~33.3 Mb. Structural comparisons to LdBPK282A1 revealed numerous rearrangements, including some reshuffling genes among polycistronic transcription units and validated by polycistronic reads from RNA sequencing. Promastigote and amastigote RNA sequencing produced 469,010 and 46,729 monocistronic reads containing a spliced-leader and a polyA tail sequences, defining 8,479 transcripts and supporting 7,415 of the 7,969 annotated protein coding genes, as well as 604 putative long non-coding RNAs. We annotated UTRs for 4,921 genes and observed that putative RNA G-quadruplexes were markedly enriched in UTRs. We also noted that 31.9% and 11.5% were expressed into multiple isoforms in promastigotes and amastigotes, respectively. Collectively, these data provide a comprehensive annotation of L. donovani genes and their UTRs and reveal widespread and stage-specific UTR length polymorphisms, and, overall, points to an important role of 3' UTR in post-transcriptional regulation in L. donovani.

Journal Article↗

PubMed2026

Translating single-cell RNA sequencing into monocyte direct leukocyte subpopulation-transcript abundance assay ratio-based biomarkers (IFI27/PSAP or IFI27/CTSS) for clinical detection of viral infection.

A rapid method for triaging febrile patients by aetiology (e.g., viral or bacterial infection) using gene expression in peripheral blood (PB) is an intensively researched area. However, gene expression in blood represents a composite sum of gene expression of all the component cell types present in the sample. As a result, numerous genes are measured in most proposed signatures. Herein, we propose a simple ratio-based biomarker (RBB) called direct leukocyte subpopulation-transcript abundance assay (DIRECT LS-TA) that recapitulates gene expressions of a single cell type in PB (i.e., monocytes). Based on single-cell RNA sequencing (scRNAseq) data and bulk expression data, IFI27 and SIGLEC1 are found as interferon-stimulated genes (ISGs) predominantly expressed by monocytes. The DIRECT LS-TA method can use a simple ratio of two genes measured in PB as an RBB to represent the target gene expression in monocytes without the need for monocyte purification. Both scRNAseq and bulk RNA sequencing datasets were used to evaluate the correlation between ISG expression in monocytes and PB, with a particular focus on monocyte expression of IFI27. An iceberg plot of bulk transcriptome data was used to identify genes that were predominantly expressed by monocytes in PB. DIRECT LS-TA RBBs of the three genes (IFI27, IFI44L and SIGLEC1) were evaluated by group-wise comparison, receiver operating characteristic and meta-analysis. In addition, the conventional interferon (IFN) score was evaluated for comparison of diagnostic performance. In viral infection datasets, DIRECT LS-TA of IFI27 (IFI27/PSAP or IFI27/CTSS) was most intensely activated (p value by t test <1e-9) and had the best area under the curve (0.94) among the three potential monocyte ISGs analysed. DIRECT LS-TA SIGLEC1 was also another monocyte biomarker but showed a lower activation (p<9e-5). IFI27/PSAP showed better diagnostic performance than the conventional IFN score. On the other hand, IFI44L was not a predominant monocyte expression gene. DIRECT LS-TA of IFI27 (IFI27/PSAP or IFI27/CTSS) measured in PB was the best biomarker of viral infection and IFN activation among ISGs predominantly expressed by monocytes. It performed even better than the conventional IFN score which required quantification of eight genes. The results suggest that DIRECT LS-TA of IFI27 is a monocyte-informative biomarker which is easy to determine in PB without the need for cell sorting.

Humans↗