It's a DoG-eat-DoG world-altered transcriptional mechanisms drive downstream-of-gene (DoG) transcript production.

The past decade has revolutionized our understanding of regulatory noncoding RNAs (ncRNAs). Among the most recently identified ncRNAs are downstream-of-gene (DoG)-containing transcripts that are produced by widespread transcriptional readthrough. The discovery of DoGs has set the stage for future studies to address many unanswered questions regarding the mechanisms that promote readthrough transcription, RNA processing, and the cellular functions of the unique transcripts. In this review, we summarize current findings regarding the biogenesis, function, and mechanisms regulating this exciting new class of RNA molecules.

[Genomics and proteomics in the research of neuromuscular diseases]. / Genomik und Proteomik in der Erforschung neuromuskulärer Erkrankungen.

Neurological diseases affect 3-5% of children and, apart from cardiovascular diseases and cancer, represent the most prominent cause of morbidity and mortality in adults and particularly in the aged population of western Europe. Neuromuscular disorders are a subgroup of neurological diseases and often have a genetic origin, which leads to familial clustering. Despite the enormous progress in the analysis of the genome, such as by sequence analysis of coding regions of deoxyribonucleic acid or even the entire deoxyribonucleic acid sequence, in approximately 50% of the patients suffering from rare forms of neurological diseases the genetic cause remains unsolved. The reasons for this limited detection rate are presented in this article. If a treatment concept is available, under certain conditions this can have an impact on the adequate and early treatment of these patients. Considering neuromuscular disorders as a paradigm, this article reports on the advantages of the inclusion of next generation sequencing analysis-based DNA investigations as an omics technology (genomics) and the advantage of the integration with protein analyses (proteomics). A special focus is on the combination of genomics and proteomics in the sense of a proteogenomic approach in the diagnostics and research of these diseases. Along this line, this article presents a proteogenomic approach in the context of a multidisciplinary project aiming towards improved diagnostic work-up and future treatment of patients with neuromuscular diseases; "NMD-GPS: gene and protein signatures as a global positioning system in patients suffering from neuromuscular diseases".

ARTDeco: automatic readthrough transcription detection.

BACKGROUND: Mounting evidence suggests several diseases and biological processes target transcription termination to misregulate gene expression. Disruption of transcription termination leads to readthrough transcription past the 3' end of genes, which can result in novel transcripts, changes in epigenetic states and altered 3D genome structure. RESULTS: We developed Automatic Readthrough Transcription Detection (ARTDeco), a tool to detect and analyze multiple features of readthrough transcription from RNA-seq and other next-generation sequencing (NGS) assays that profile transcriptional activity. ARTDeco robustly quantifies the global severity of readthrough phenotypes, and reliably identifies individual genes that fail to terminate (readthrough genes), are aberrantly transcribed due to upstream termination failure (read-in genes), and novel transcripts created as a result of readthrough (downstream of gene or DoG transcripts). We used ARTDeco to characterize readthrough transcription observed during influenza A virus (IAV) infection, validating its specificity and sensitivity by comparing its performance in samples infected with a mutant virus that fails to block transcription termination. We verify ARTDeco's ability to detect readthrough as well as identify read-in genes from different experimental assays across multiple experimental systems with known defects in transcriptional termination, and show how these results can be leveraged to improve the interpretation of gene expression and downstream analysis. Applying ARTDeco to a gene expression data set from IAV-infected monocytes from different donors, we find strong evidence that read-in gene-associated expression quantitative trait loci (eQTLs) likely regulate genes upstream of read-in genes. This indicates that taking readthrough transcription into account is important for the interpretation of eQTLs in systems where transcription termination is blocked. CONCLUSIONS: ARTDeco aids researchers investigating readthrough transcription in a variety of systems and contexts.

Characterization of blaCTX-M-27/F1:A2:B20 Plasmids Harbored by Escherichia coli Sequence Type 131 Sublineage C1/H30R Isolates Spreading among Elderly Japanese in Nonacute-Care Settings.

We characterized 29 blaCTX-M-27-harboring plasmids of Escherichia coli sequence type 131 (ST131) sublineage C1/H30R isolates from healthy individuals and long-term-care facility (LTCF) residents. Most (27/29) plasmids were of the FIA, FIB, and FII multireplicon type with the same plasmid multilocus sequence typing (pMLST). Several plasmids (7/23) from LTCF residents harbored only blaCTX-M-27 as the resistance gene; however, their fundamental structures were very similar to those of previously isolated blaCTX-M-27/F1:A2:B20 plasmids, suggesting their prevalence as a newly arising public health concern.

Bioinformatics Tools for Genome-Wide Epigenetic Research.

Epigenetics play a central role in the regulation of many important cellular processes, and dysregulations at the epigenetic level could be the source of serious pathologies, such as neurological disorders affecting brain development, neurodegeneration, and intellectual disability. Despite significant technological advances for epigenetic profiling, there is still a need for a systematic understanding of how epigenetics shapes cellular circuitry, and disease pathogenesis. The development of accurate computational approaches for analyzing complex epigenetic profiles is essential for disentangling the mechanisms underlying cellular development, and the intricate interaction networks determining and sensing chromatin modifications and DNA methylation to control gene expression. In this chapter, we review the recent advances in the field of "computational epigenetics," including computational methods for processing different types of epigenetic data, prediction of chromatin states, and study of protein dynamics. We also discuss how "computational epigenetics" has complemented the fast growth in the generation of epigenetic data for uncovering the main differences and similarities at the epigenetic level between individuals and the mechanisms underlying disease onset and progression.

VarElect: the phenotype-based variation prioritizer of the GeneCards Suite.

BACKGROUND: Next generation sequencing (NGS) provides a key technology for deciphering the genetic underpinnings of human diseases. Typical NGS analyses of a patient depict tens of thousands non-reference coding variants, but only one or very few are expected to be significant for the relevant disorder. In a filtering stage, one employs family segregation, rarity in the population, predicted protein impact and evolutionary conservation as a means for shortening the variation list. However, narrowing down further towards culprit disease genes usually entails laborious seeking of gene-phenotype relationships, consulting numerous separate databases. Thus, a major challenge is to transition from the few hundred shortlisted genes to the most viable disease-causing candidates. RESULTS: We describe a novel tool, VarElect ( http://ve.genecards.org ), a comprehensive phenotype-dependent variant/gene prioritizer, based on the widely-used GeneCards, which helps rapidly identify causal mutations with extensive evidence. The GeneCards suite offers an effective and speedy alternative, whereby >120 gene-centric automatically-mined data sources are jointly available for the task. VarElect cashes on this wealth of information, as well as on GeneCards' powerful free-text Boolean search and scoring capabilities, proficiently matching variant-containing genes to submitted disease/symptom keywords. The tool also leverages the rich disease and pathway information of MalaCards, the human disease database, and PathCards, the unified pathway (SuperPaths) database, both within the GeneCards Suite. The VarElect algorithm infers direct as well as indirect links between genes and phenotypes, the latter benefitting from GeneCards' diverse gene-to-gene data links in GenesLikeMe. Finally, our tool offers an extensive gene-phenotype evidence portrayal ("MiniCards") and hyperlinks to the parent databases. CONCLUSIONS: We demonstrate that VarElect compares favorably with several often-used NGS phenotyping tools, thus providing a robust facility for ranking genes, pointing out their likelihood to be related to a patient's disease. VarElect's capacity to automatically process numerous NGS cases, either in stand-alone format or in VCF-analyzer mode (TGex and VarAnnot), is indispensable for emerging clinical projects that involve thousands of whole exome/genome NGS analyses.

Identification of Long Noncoding RNAs Expression Profiles Between Gallstone and Gallbladder Cancer Using Next-Generation Sequencing Analysis.

Background: Gallstone disease (GS) is an important risk factor for Gallbladder cancer (GBC). However, the mechanisms of the progression of GS to GBC remain unclear. Long non-coding RNA (lncRNA), modulates DNA/RNA/proteins at epigenetic, pre-transcriptional, transcriptional and posttranscriptional levels, and plays a potential therapeutic role in various diseases. This study aims to identify lncRNAs that have a potential impact on GS-promoted GBC progression. Methods and Results: Six GBC patients without GS, six normal gallbladder tissues, nine gallstones and nine GBC patients with GS were admitted to our hospital. The next-generation RNA-sequencing was performed to analyze differentially expressed (DE) lncRNA and messenger RNA (mRNA) in four groups. Then overlapping and specific molecular signatures were analyzed. We identified 29 co-DEGs and 500 co-DElncRNAs related to gallstone or GBC. The intersection and concatenation of co-DEGs and co-DElncRNA functionally involved in focal adhesion, Transcriptional misregulation in cancers, Protein digestion and absorption, and ECM-receptor interaction signaling pathways may contribute to the development of gallbladder cancer. Further exploration is necessary for early diagnosis and the potential treatment of GBC. FXYD2, MPZL1 and PAH were observed in both co-DEGs and co-DElncRNA and validated by qRT-PCR. Conclusion: Our data identified a series of DEGs and DElncRNAs, which were involved in the progression of GBC and GS-related metabolism pathways. Compared to GBC, the GS profile was more similar to para-tumor tissues in transcriptome level and lower risk of cancer. Further exploration is necessary from GBC patients with different periods of follow-up gallstone.

[Corrigendum] Next­generation sequencing analysis reveals that MTH­3, a novel curcuminoid derivative, suppresses the invasion of MDA­MB­231 triple­negative breast adenocarcinoma cells.

Subsequently to the publication of the article, an interested reader drew to the authors' attention that, in Fig. 2A on p. 5, the 'Control  (24 h)' and 'MTH­3 (1 µM; 24 h)' data panels contained partially overlapping data, such that they appeared to have been derived from the same original source. The authors have examined their original data, and realized that this error arose inadvertently as a consequence of having compiled this figure incorrectly. The revised version of Fig. 2, featuring the data from one of the repeated experiments in Fig. 2A, is shown below. The revised data shown for this figure do not affect the overall conclusions reported in the paper. The authors apologize to the Editor of Oncology Reports and to the readership for any inconvenience caused. [Oncology Reports 46: 133, 2021; DOI: 10.3892/or.2021.8084].

Investigation of the molecular genetic causes of non-syndromic primary ovarian insufficiency by next generation sequencing analysis.

Objective: The aim of this study is to investigate the molecular genetic causes of non-syndromic primary ovarian insufficiency (POI) cases with the gene panel basedon next generation sequencing analysis and to establish the relationship between genotype and phenotype. Materials and methods: Twenty three cases aged 14-40 years followed up with POI were included. Patients with a karyotype of 46, XX, primary or secondary amenorrhea before the age of 40, with elevated FSH (>40 IU/mL) and low AMH levels (<0.03 ng/mL) were included in the study. Molecular genetic analyzes were performed by the next generation sequencing analysis method targeted with the TruSight TM Exome panel. Results: Median age of the cases was 17.8 (14.0-24.3) years, and 12 (52%) cases admitted before the age of 18. Fifteen (65%) patients had consanguineous parents. In2 (8.6%) cases, variants detected were in genes that have been previously proven to cause POI. One was homozygous variant in FIGLA gene and the other was homozygous variant in PSMC3IP gene. Heterozygous variants were detected in PROK2, WDR11 and CHD7 associated with hypogonadotropic hypogonadism, but these variants are insufficient to contribute to the POI phenotype. Conclusion: Genetic panels based on next generation sequencing analysis technologies can be used to determine the molecular genetic diagnosis of POI, which has a highly heterogeneous genetic basis.

Transcriptomic analysis of stem cells from chorionic villi uncovers the impact of chromosomes 2, 6 and 22 in the clinical manifestations of Down syndrome.

BACKGROUND: Down syndrome (DS) clinical multisystem condition is generally considered the result of a genetic imbalance generated by the extra copy of chromosome 21. Recent discoveries, however, demonstrate that the molecular mechanisms activated in DS compared to euploid individuals are more complex than previously thought. Here, we utilize mesenchymal stem cells from chorionic villi (CV) to uncover the role of comprehensive functional genomics-based understanding of DS complexity. METHODS: Next-generation sequencing coupled with bioinformatic analysis was performed on CV obtained from women carrying fetuses with DS (DS-CV) to reveal specific genome-wide transcriptional changes compared to their euploid counterparts. Functional assays were carried out to confirm the biological processes identified as enriched in DS-CV compared to CV (i.e., cell cycle, proliferation features, immunosuppression and ROS production). RESULTS: Genes located on chromosomes other than the canonical 21 (Ch. 2, 6 and 22) are responsible for the impairment of life-essential pathways, including cell cycle regulation, innate immune response and reaction to external stimuli were found to be differentially expressed in DS-CV. Experimental validation confirmed the key role of the biological pathways regulated by those genes in the etiology of such a multisystem condition. CONCLUSIONS: NGS dataset generated in this study highlights the compromised functionality in the proliferative rate and in the innate response of DS-associated clinical conditions and identifies DS-CV as suitable tools for the development of specifically tailored, personalized intervention modalities.

Molecular diagnosis in patients with monogenic diabetes mellitus, and detection of a novel candidate gene.

AIM: We aimed to investigate molecular genetic basis of monogenic diabetes (DM) and novel responsible candidate genes with targeted Next Generation Sequencing (NGS) and Whole Exome Sequencing (WES). METHODS: A hundred cases presenting with clinical findings and a family history of monogenic DM were included in the study. Molecular analysis was performed using an NGS panel including 14 genes. Following targeted NGS, WES was planned in cases in whom no variant was detected. RESULTS: Thirty different disease-causing variants in seven different genes were detected in thirty-five (35 %) cases with targeted NGS approach. Most common pathogenic variant was found in GCK gene in 25 (25 %) cases. Four different variants were detected in 4 (4 %) patients in ABCC8 gene. In 45 of 65 cases; WES analyses were done. A heterozygous c.2635C > T(p.Gln879Ter) variant was detected in IFIH1 gene in a patient with incidental hyperglycemia. In the segregation analysis affected mother was shown to be heterozygous for the same variant. CONCLUSION: Molecular etiology was determined in 35 % cases with the NGS targeted panel. Seventeen novel variants in monogenic DM genes have been identified. A candidate gene determined by WES analysis in a case that could not be diagnosed with NGS panel in this study.

Genomic alterations associated with pseudoprogression and hyperprogressive disease during anti-PD1 treatment for advanced non-small-cell lung cancer.

Introduction: This study aimed to elucidate the relationship between dynamic genomic mutation alteration and pseudoprogression (PsPD)/hyperprogressive disease (HPD) in immunotherapy-treated advanced non-small-cell lung cancer (NSCLC), to provide clinical evidence for identifying and distinguishing between PsPD and HPD. Method: Patients with advanced NSCLC who were treated with anti-PD1 were enrolled. Whole blood was collected at baseline and post image progression. Serum was separated and sequenced using 425-panel next-generation sequencing analysis (NGS). Results: NGS revealed that not only single gene mutations were associated with PsPD/HPD before treatment, dynamic monitoring of the whole-blood genome mutation spectrum also varied greatly. Mutational burden, allele frequency%, and relative circulating tumor DNA abundance indicated that the fold change after image progression was much higher in the HPD group. Discussion: The gene mutation profiles of PsPD and HPD not only differed before treatment, but higher genome mutation spectrum post image progression indicated true disease progression in patients with HPD. This suggests that dynamic whole-genome mutation profile monitoring as NGS can distinguish PsPD from HPD more effectively than single gene detection, providing a novel method for guiding clinical immune treatment.

A look towards the clonal origin of metastatic pulmonary carcinosarcoma: Report of a patient with an unexpected long-term survival.

INTRODUCTION: Pulmonary carcinosarcoma is a rare histological subtype of non-small cell lung cancer, defined by the combination of epithelial and mesenchimal elements. Prognosis is usually dismal, with a median survival of about 6 months. The use of immunotherapy by blockade of PD1/PD-L1 immune checkpoint signaling has been shown to improve patients' survival. However, local aggressiveness and distant metastases are frequent. Spread to the gastrointestinal tract is seldom reported. The genetic landscape of the disease has only recently begun to emerge, pointing at TP53, KRAS, EGFR and MET as the most common mutated genes. CASE DESCRIPTION: We describe the case of a metastatic patient with 37 months overall survival, treated by an aggressive multimodal approach combining surgery, chemotherapy, radiotherapy and immunotherapy. To shed new light on the molecular basis for sarcomatoid component in lung carcinoma, we performed next generation sequencing analysis of the squamous and sarcomatoid component by the two sites. We demonstrated a clonal origin and hypermutability of the sarcomatous elements that may account for the good response to immunotherapy. Moreover, we identified some mutations involving TP53 and EGFR genes, targetable by already available drugs. CONCLUSIONS: We depicted a model of how a squamous cell carcinoma can differentiate during its natural history into sub-clonal populations with different features and may ultimately result in a neoplasm (i.e. pulmonary carcinosarcoma) showing clonal heterogeneity. Our data might contribute to a better understanding of the pathogenesis and molecular mechanisms of this rare tumor and open new ways for a more tailored approach.

Clinical and Genetic Analysis of a Patient with CMT4J.

We report the clinical and genetic analysis of a patient with a rare form of an autosomal recessive genetic neuropathy, Charcot Marie Tooth (CMT) disease type 4J. She presented at age 62 years with signs and symptoms consistent with a mild neuropathy. The onset of symptoms began approximately ten years earlier. Electrophysiological testing confirmed a demyelinating neuropathy and a comprehensive neuropathy screening for common causes of neuropathy was unrevealing. She underwent commercial whole exome sequencing, analyzing more than eighty genes known to cause neuropathy. Two mutations were detected, c.122T > C, p.Ile41Thr and c.2247dupC, p.Ser750GlnX10 in the FIG4 gene. The p.Ile41Thr mutation, which is paternally inherited, is a recurrent mutation reported in a number of unrelated families of European descent. The patient's father, also of European descent, provides further evidence supporting a founder effect for this mutation. In most patients carrying the p.Ile41Thr mutation, the neuropathy, unlike our patient, is often severe with early onset. The second mutation, c.2247dupC, p.Ser750GlnX10 is maternally inherited and not previously reported. Furthermore, based upon our protein modeling analysis, c.2247dupC is disease producing, representing a novel pathogenic mutation. Our study of this patient expands the clinical and genetic spectrum of patients with CMT 4J.

Comprehensive Analyses of the Bacterial Population in Non-Healing Claw Lesions of Dairy Cattle.

Non-healing claw lesions (NHCLs) are a newly characterized disorder affecting the deep dermis of the hoof in dairy cattle. Although NHCLs are thought to be associated with bovine digital dermatitis (BDD), their precise etiology is not yet understood. To investigate the bacterial populations present in each type of NHCL (toe necrosis: TN, non-healing white line disease: nhWLD, and a non-healing sole ulcer: nhSU), and the newly added entity non-healing verrucous-like lesions (nhVLL), 16S rRNA-based metagenomic analysis with next-generation sequencing (NGS) was employed. Twelve cases of NHCLs (3 TN, 3 nhWLD, 4 nhSU, and 2 nhVLL) were collected from five dairy farms in two prefectures in Japan. Three samples of healthy hoof dermis collected from two farms and a slaughterhouse were used as controls. Furthermore, culture-dependent and -independent approaches were conducted for detecting Treponema species and Fusobacterium necrophorum. As reported in BDD, Treponema species and F. necrophorum were detected frequently from NHCLs by PCR and immunohistochemistry, but NGS showed that these bacterial genera were not predominant in NHCLs. The predominant bacterial genera in NHCLs differed among the lesions examined, suggesting that Treponema species present predominantly in BDD were not predominant in NHCLs and that the bacterial population in NHCLs may vary among individual cattle and/or farms.

Next­generation sequencing analysis reveals that MTH­3, a novel curcuminoid derivative, suppresses the invasion of MDA­MB­231 triple­negative breast adenocarcinoma cells.

Triple­negative breast cancer (TNBC) behaves aggressively in the invasive and metastatic states. Our research group recently developed a novel curcumin derivative, (1E,3Z,6E)-3-hydroxy-5-oxohepta-1,3,6-triene-1,7-diyl)bis(2­methoxy-4,1­phenylene)bis(3-hydroxy2-hydroxymethyl)-2­methyl propanoate (MTH­3), and previous studies showed that MTH­3 inhibits TNBC proliferation and induces apoptosis in vitro and in vivo with a superior bioavailability and absorption than curcumin. In the present study, the effects of MTH­3 on TNBC cell invasion were examined using various assays and gelatin zymography, and western blot analysis. Treatment with MTH­3 inhibited MDA­MB­231 cell invasion and migration, as shown by Transwell assay, 3D spheroid invasion assay, and wound healing assay. The results of the gelatin zymography experiments revealed that MTH­3 decreased matrix metalloproteinase­9 activity. The potential signaling pathways were revealed by next­generation sequencing analysis, antibody microarray analysis and western blot analysis. In conclusion, the results of the present study show that, MTH­3 inhibited tumor cell invasion through the MAPK/ERK/AKT signaling pathway and cell cycle regulatory cascade, providing significant information about the potential molecular mechanisms of the effects of MTH­3 on TNBC.

Enrichment of a mixed microbial culture of PHA-storing microorganisms by using fermented hardwood spent sulfite liquor.

Pulp and paper factories produce several residues that can be explored and valorized through polyhydroxyalkanoate (PHA) production via a three-step process. The objective of this work was focused on the selection step. Acidified hardwood spent sulfite liquor (HSSL), a fermented waste stream from a pulp and paper factory, was used to select a mixed microbial culture (MMC) in a sequencing batch reactor (SBR) operated for 156 days under different operational conditions. The MMC adapted to the imposed conditions, revealing its robustness whenever the operational parameters were changed. Feast-to-Famine ratio was kept below or equal to 0.2, with constant production of a copolymer of P(3HB-co-3 HV), and with storage contents values over 30 %. Changes in the operational conditions, namely cycle length, and organic load rate (OLR), successfully led to the selection of an MMC with a stable accumulation capacity and an increased biomass concentration. Next Generation Sequencing analysis was performed on samples collected during the SBR operational period. The analysis of the microbial composition of the MMC showed a rise in PHA-accumulating bacteria over time. Acidovorax and Comamonas species were found mainly to drive the PHA storage process during the first two periods of operation. After an increase in the OLR, in the last period, a shift towards Comamonas dominance occurred, suggesting a higher tolerance to the inhibitory compounds of the HSSL for this genus.

Characterization of eubacterial communities by Denaturing Gradient Gel Electrophoresis (DGGE) and Next Generation Sequencing (NGS) in a desulfurization biotrickling filter using progressive changes of nitrate and nitrite as final electron acceptors.

Anoxic biotrickling filters (BTFs) represent a technology with high H2S elimination capacity and removal efficiencies widely studied for biogas desulfurization. Three changes in the final electron acceptors were made using nitrate and nitrite during an operating period of 520 days. The stability and performance of the anoxic BTF were maintained when a significant perturbation was applied to the system that involved the progressive change of nitrate to nitrite and vice versa. Here the impact of electron acceptor changes on the microbial community was characterized by denaturing gel gradient electrophoresis (DGGE) and next generation sequencing (NGS). Both platforms revealed that the community underwent changes during the perturbations but was resilient because the removal capacity did not significantly change. Proteobacteria and Bacteroidetes were the main Phyla and Sulfurimonas and Thiobacillus the main nitrate-reducing sulfide-oxidizing bacteria (NR-SOB) genera involved in the biodesulfurization process.

Investigation of the molecular genetic causes of non-syndromic primary ovarian ınsufficiency by next generation sequencing analysis

ABSTRACT Objective: The aim of this study is to investigate the molecular genetic causes of non-syndromic primary ovarian insufficiency (POI) cases with the gene panel based on next generation sequencing analysis and to establish the relationship between genotype and phenotype. Subjects and methods: Twenty three cases aged 14-40 years followed up with POI were included. Patients with a karyotype of 46, XX, primary or secondary amenorrhea before the age of 40, with elevated FSH (>40 IU/mL) and low AMH levels (<0.03 ng/mL) were included in the study. Molecular genetic analyzes were performed by the next generation sequencing analysis method targeted with the TruSightTM Exome panel. Results: Median age of the cases was 17.8 (14.0-24.3) years, and 12 (52%) cases admitted before the age of 18. Fifteen (65%) patients had consanguineous parents. In 2 (8.6%) cases, variants detected were in genes that have been previously proven to cause POI. One was homozygous variant in FIGLA gene and the other was homozygous variant in PSMC3IP gene. Heterozygous variants were detected in PROK2, WDR11 and CHD7 associated with hypogonadotropic hypogonadism, but these variants are insufficient to contribute to the POI phenotype. Conclusion: Genetic panels based on next generation sequencing analysis technologies can be used to determine the molecular genetic diagnosis of POI, which has a highly heterogeneous genetic basis.

Screening of DNA Signaling Aptamer from Multiple Candidates Obtained from SELEX with Next-generation Sequencing.

Here, we demonstrated a strategy for developing signaling aptamers, based on screening of signaling aptamers from multiple aptamer candidates obtained by SELEX with next generation sequencing. Among aptamer candidates labelled by 6-carboxyfluorescein and quencher at both end termini, there is the possibility of discovering a potent signaling aptamer. In this study, we discovered DNA signaling aptamers against VEGFR-1. This strategy has the potential for signaling aptamer discovery without the extremely laborious task of optimization of oligodeoxynucleotide modifications.