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Publications

 To see one of Swift’s many publication references, simply expand each accordion section below.

Rhodoliths holobionts in a changing ocean: host-microbes interactions mediate coralline algae resilience under ocean acidification

BMC Genomics 2018. 19:701 https://doi.org/10.1186/s12864-018-5064-4

Life in the ocean will increasingly have to contend with a complex matrix of concurrent shifts in environmental properties that impact their physiology and control their life histories. Rhodoliths are coralline red algae (Corallinales, Rhodophyta) that are photosynthesizers, calcifiers, and ecosystem engineers and therefore represent important targets for ocean acidification (OA) research. Here, we exposed live rhodoliths to near-future OA conditions to investigate responses in their photosynthetic capacity, calcium carbonate production, and associated microbiome using carbon uptake, decalcification assays, and whole genome shotgun sequencing metagenomic analysis, respectively…more.

The Mcm2-Ctf4-Polα Axis Facilitates Parental Histone H3-H4 Transfer to Lagging Strands

Molecular Cell (2018). Science Direct. https://doi.org/10.1016/j.molcel.2018.09.001

Although essential for epigenetic inheritance, the transfer of parental histone (H3-H4)2tetramers that contain epigenetic modifications to replicating DNA strands is poorly understood. Here, we show that the Mcm2-Ctf4-Polα axis facilitates the transfer of parental (H3-H4)2 tetramers to lagging-strand DNA at replication forks. Mutating the conserved histone-binding domain of the Mcm2 subunit of the CMG (Cdc45-MCM-GINS) DNA helicase, which translocates along the leading-strand template, results in a marked enrichment of parental (H3-H4)2 on leading strand, due to the impairment of the transfer of parental (H3-H4)2 to lagging strands. Similar effects are observed in Ctf4 and Polα primase mutants that disrupt the connection of the CMG helicase to Polα that resides on lagging-strand template. Our results support a model whereby parental (H3-H4)2 complexes displaced from nucleosomes by DNA unwinding at replication forks are transferred by the CMG-Ctf4-Polα complex to lagging-strand DNA for nucleosome assembly at the original location…more.

T follicular helper cells restricted by IRF8 contribute to T cell-mediated inflammation

Journal of Autoimmunity (2018). Science Direct. https://doi.org/10.1016/j.jaut.2018.09.001

The follicular helper T cell (TFH) are established regulators of germinal center (GC) B cells, whether TFH have pathogenic potential independent of B cells is unknown. Based on in vitroTFH cell differentiation, in vivo T cell transfer animal colitis model, and intestinal tissues of inflammatory bowel disease (IBD) patients, TFH and its functions in colitis development were analyzed by FACS, ChIP, ChIP-sequencing, WB, ELISA and PCR. Herein we demonstrate that intestinal tissues of patients and colon tissues obtained from Rag1−/− recipients of naïve CD4+ T cells with colitis, each over-express TFH-associated gene products. Adoptive transfer of naïve Bcl6−/− CD4+ T cells into Rag1−/− recipient mice abrogated development of colitis and limited TFH differentiation in vivo, demonstrating a mechanistic link. In contrast, T cell deficiency of interferon regulatory factor 8 (IRF8) resulted in augmentation of TFH induction in vitro and in vivomore.

Detection of Viruses in Clinical Samples Using Metagenomic Sequencing and Targeted Sequence Capture

Journal of Clinical Microbiology (2018), DOI: 10.1128/JCM.01123-18

Metagenomic shotgun sequencing (MSS) is a revolutionary approach to viral diagnostic testing that allows simultaneous detection of a broad range of viruses, detailed taxonomic assignment, and detection of mutations associated with antiviral drug resistance. To enhance sensitivity for virus detection, we previously developed ViroCap, a targeted sequence capture panel designed to enrich nucleic acid from a comprehensive set of eukaryotic viruses prior to sequencing. To demonstrate the utility of MSS with targeted sequence capture for detecting clinically important viruses and characterizing clinically important viral features, we used ViroCap to analyze clinical samples from a diagnostic virology laboratory containing a broad range of medically relevant viruses. From 26 samples, MSS with ViroCap detected all of the expected viruses and 30 additional viruses…more.

Necroptosis microenvironment directs lineage commitment in liver cancer

Nature (2018), https://doi.org/10.1038/s41586-018-0519-y

Primary liver cancer represents a major health problem. It comprises hepatocellular carcinoma (HCC) and intrahepatic cholangiocarcinoma (ICC), which differ markedly with regards to their morphology, metastatic potential and responses to therapy. However, the regulatory molecules and tissue context that commit transformed hepatic cells towards HCC or ICC are largely unknown. Here we show that the hepatic microenvironment epigenetically shapes lineage commitment in mosaic mouse models of liver tumorigenesis. Whereas a necroptosis-associated hepatic cytokine microenvironment determines ICC outgrowth from oncogenically transformed hepatocytes, hepatocytes containing identical oncogenic drivers give rise to HCC if they are surrounded by apoptotic hepatocytes….more.

Genomic Evaluation of Multiparametric Magnetic Resonance Imaging-visible and -nonvisible Lesions in Clinically Localised Prostate Cancer

Parry MA, et al. (2018), https://doi.org/10.1016/j.euo.2018.08.005

Background: The prostate cancer (PCa) diagnostic pathway is undergoing a radical change with the introduction of multiparametric magnetic resonance imaging (mpMRI), genomic testing, and different prostate biopsy techniques. It has been proposed that these tests should be used in a sequential manner to optimise risk stratification. Objective: To characterise the genomic, epigenomic, and transcriptomic features of mpMRI-visible and -nonvisible PCa in clinically localised disease. Design, setting, and participants: Multicore analysis of fresh prostate tissue sampled immediately after radical prostatectomy was performed for intermediate- to high-risk PCa. Intervention: Low-pass whole-genome, exome, methylation, and transcriptome profiling of patient tissue cores taken from microscopically benign and cancerous…more.

Epigenetic Profiling of Active Enhancers in Mouse Retinal Ganglion Cells

Xing, Jian (2018). University of Connecticut. https://opencommons.uconn.edu/gs_theses/1268

Retinal ganglion cells (RGCs) are projection neurons of the eye, which process and pass visual information collected in the eyes to the brain. However, epigenetic regulation of RGC fate specification remains poorly understood, in large part due to the technical challenges associated with purifying RGCs, which comprise only 1% of all retinal cells, and performing ChIP-seq profiling on a small number of cells. To overcome these limitations, we have purified RGCs from multiple mice by immunoppanning for Thy1, a surface marker on RGCs, and analyzed pulled chromatin by ChIP-seq for histone 3 acetylated at K27 (H3K27ac), which is an epigenetic marker of active (i.e. accessible) enhancers. We also utilized recently developed ChIP-seq Kit for processing small cell number samples and ultra low chromatin input (truChIP Ultra Low Cell Chromatin Shearing Kit, Covaris; Zymo-Spin ChIP Kit, Zymo Research; and Accel-NGS 2S Plus DNA Library Kit, Swift Biosciences)…more. 

Genome-wide meta-analysis and replication studies in multiple ethnicities identify novel adolescent idiopathic scoliosis susceptibility loci

 Oxford University Press. Human Molecular Genetics. 24 August 2018. doi: 10.1093/hmg/ddy306

Adolescent idiopathic scoliosis (AIS) is the most common musculoskeletal disorder of childhood development. The genetic architecture of AIS is complex, and the great majority of risk factors are undiscovered. To identify new AIS susceptibility loci we conducted the first genome-wide meta-analysis of AIS GWAS, including 7,956 cases and 88,459 controls from three ancestral groups. Three novel loci that surpassed genome-wide significance were uncovered in intragenic regions of the CDH13 (P-value_rs4513093 = 1.7E-15), ABO (P-value_ rs687621 = 7.3E-10) and SOX6 (P-value_rs1455114= 2.98E-08) genes…more.

Genome Sequences of 34 Shiga ToxinProducing Escherichia coli Isolates from Swine and Other Sources

Genome Announcements. 2017 November. doi: 10.1128/genomeA.01214-17.

Shiga toxin-producing Escherichia coli (STEC) bacteria are foodborne pathogens that can be carried by various animals. The swine STEC population is partially composed of host-specific strains that are often not well characterized. In this work, the genome sequences of a number of swine STEC strains are presented…more »

Differences between germline and somatic mutation rates in humans and mice

Nat Commun. 2017 May 9;8:15183. doi: 10.1038/ncomms15183.

The germline mutation rate has been extensively studied and has been found to vary greatly between species, but much less is known about the somatic mutation rate in multicellular organisms, which remains very difficult to determine. Presented is data on somatic mutation rates in mice and humans, obtained by sequencing single cells and clones derived from primary fibroblasts, which allows us to make the first direct comparison with germline mutation rates in these two species. The results indicate that the somatic mutation rate is almost two orders of magnitude higher than the germline mutation rate…more »

Zika virus evolution and spread in the Americas

bioRxiv preprint first posted online Feb. 18, 2017; doi: http://dx.doi.org/10.1101/109348.

Although the recent Zika virus (ZIKV) epidemic in the Americas and its link to birth defects have attracted a great deal of attention, much remains unknown about ZIKV disease epidemiology and ZIKV evolution, in part owing to a lack of genomic data. Here we address this gap in knowledge by using multiple sequencing approaches to generate 110 ZIKV genomes from clinical and mosquito samples from 10 countries and territories, greatly expanding the observed viral genetic diversity from this outbreak…more »

Accurate identification of single-nucleotide variants in whole-genome-amplified single cells

Nat Methods. 2017 May;14(5):491-493. doi: 10.1038/nmeth.4227. Epub 2017 Mar 20.

Mutation analysis in single-cell genomes is prone to artifacts associated with cell lysis and whole-genome amplification. Here we addressed these issues by developing single-cell multiple displacement amplification (SCMDA) and a general-purpose single-cell-variant caller, SCcaller. By comparing SCMDA-amplified single cells with unamplified clones from the same population, we validated the procedure as a firm foundation for standardized somatic-mutation analysis in single-cell genomics….more »

Cell-free DNA and next-generation sequencing in the service of personalized medicine for lung cancer

Oncotarget. 2016 Aug 30. DOI: 10.18632/oncotarget.11717

Personalized medicine has emerged as the future of cancer care to ensure that patients receive individualized treatment specific to their needs. In order to provide such care, molecular techniques that enable oncologists to diagnose, treat, and monitor tumors are necessary. In the field of lung cancer, cell free DNA (cfDNA) shows great potential as a less invasive liquid biopsy technique, and next-generation sequencing (NGS) is a promising tool for analysis of tumor mutations…. more »

Vasculogenic mimicry in small cell lung cancer

Nat Commun. 2016 Nov 9;7:13322.

Small cell lung cancer (SCLC) is characterized by prevalent circulating tumour cells (CTCs), early metastasis and poor prognosis. We show that SCLC patients (37/38) have rare CTC subpopulations co-expressing vascular endothelial-cadherin (VE-cadherin)…. more »

A comparative study of ChIP-seq sequencing library preparation methods

BMC Genomics (2016) 17:816.

ChIP-seq is the primary technique used to investigate genome-wide protein-DNA interactions. As part of this procedure, immunoprecipitated DNA must undergo “library preparation” to enable subsequent high-throughput sequencing…. more »

Cell cycle progression in Caulobacter requires a nucleoid-associated protein with high AT sequence recognition

Proc Natl Acad Sci U S A. 2016 Sep 19. pii: 201612579.

In all organisms, morphological and functional diversity is the product of cell type-specific genetic programs. Asymmetric cell division in Caulobacter yields daughter cells that differ functionally due to the differential read-out of their genomes…. more »

Distinct sub-clonal tumour responses to therapy revealed by circulating cell-free DNA

Ann Oncol (2016). doi: 10.1093/annonc/mdw278. First published online: August 8, 2016.

The application of precision medicine in oncology requires in-depth characterisation of a patient’s tumours and the dynamics of their responses to treatment. We used next generation sequencing of circulating cell-free DNA to monitor the response of a KIT p.L576P-mutant metastatic vaginal mucosal melanoma to sequential targeted, immuno- and chemotherapy…. more »

Digital sorting of pure cell populations enables unambiguous genetic analysis of heterogeneous formalin-fixed paraffin-embedded tumors by next generation sequencing

Sci Rep. 2016 Feb 11;6:20944. doi: 10.1038/srep20944.

Precision medicine in oncology requires an accurate characterization of a tumor molecular profile for patient stratification. Though targeted deep sequencing is an effective tool to detect the presence of somatic sequence variants, a significant number of patient specimens do not meet the requirements needed for routine clinical application… more »

Application of sequencing, liquid biopsies, and patient-derived xenografts for personalized medicine in melanoma

Cancer Discov. 2016 Mar;6(3):286-99. doi: 10.1158/2159-8290. CD-15- 1336. Epub 2015 Dec 29.

Targeted therapies and immunotherapies have transformed melanoma care, extending median survival from ∼9 to over 25 months, but nevertheless most patients still die of their disease. The aim of precision medicine is to tailor care for individual patients and improve outcomes… more »

Genome sequence of Klebsiella pneumoniae urinary tract isolate Top52

Genome Announc. 2(4):e00668-14. doi:10.1128/genomeA.00668-14.

Klebsiella pneumoniae is a significant cause of nosocomial infections, including ventilator-associated pneumonias and catheter-associated urinary tract infections. K. pneumoniae strain TOP52 #1721 (Top52) was isolated from a woman presenting with acute cystitis and subsequently characterized using various murine models of infection… more »

Genome sequence of Klebsiella pneumoniae respiratory isolate IA565

Genome Announc. 2(5):e00896-14. doi:10.1128/genomeA.00896-14.

Klebsiella pneumoniae is a clinically significant opportunistic bacterial pathogen as well as a normal member of the human microbiota. K. pneumoniae strain IA565 was isolated from a tracheal aspirate at the University of Iowa Hospitals and Clinics… more »

Analysis of solid tumor mutation profiles in liquid biopsy

Cancer Medicine published by John Wiley & Sons Ltd. 2018. DOI: 10.1002/cam4.1791

Liquid biopsy is increasingly gaining traction as an alternative to invasive solid tumor biopsies for prognosis, treatment decisions, and disease monitoring. Matched tumor‐plasma samples were collected from 180 patients across different cancers with >90% of the samples below Stage IIIB. Tumors were profiled using next‐generation sequencing (NGS) or quantitative PCR (qPCR), and the mutation status was queried in the matched plasma using digital platforms such as droplet digital PCR (ddCPR) or NGS for concordance. Tumor‐plasma concordance of 82% and 32% was observed in advanced (Stage IIB and above) and early (Stage I to Stage IIA) stage samples, respectively. Interestingly, the overall survival outcomes correlated to presurgical/ at‐biopsy ctDNA levels. Baseline ctDNA stratified patients into three categories: (a) high ctDNA correlated with poor survival outcome, (b) undetectable ctDNA with good outcome, and (c) low ctDNA whose outcome was ambiguous. ctDNA could be a powerful tool for therapy decisions and patient management in a large number of cancers across a variety of stages…more.

Distinct clinical and pathological features of melorheostosis associated with somatic MAP2K1 mutations

Journal of Bone and Mineral Research. 23 August 2018. doi: [ 10.1002/jbmr.3577]

Melorheostosis is a rare hyperostotic disease of the long bones classically characterized by a “dripping candle‐wax” radiographic appearance. We recently described somatic activating mutations in MAP2K1 as a cause of melorheostosis. Here, we report distinguishing characteristics of patients with MAP2K1‐positive melorheostosis. Fifteen unrelated patients with radiographic appearance of melorheostosis underwent paired biopsies of affected and unaffected bone for whole exome sequencing, histology and cell culture…more.

Biotinylated Amplicon Sequencing: A method for preserving DNA samples of limited quantity

(2018), Biotinylated Amplicon Sequencing: A method for preserving DNA samples of limited quantity. ScienceDirect. https://doi.org/10.1016/j.plabm.2018.e00108

Genomic testing is often limited by the exhaustible nature of human tissue and blood samples. Here we describe Biotinylated Amplicon Sequencing (BAmSeq), a method that allows for the creation of PCR amplicon based next-generation sequencing (NGS) libraries while retaining the original source DNA. Designs and Methods: Biotinylated primers for different loci were designed to create NGS libraries using human genomic DNA from cell lines, plasma, and formalin-fixed paraffin embedded (FFPE) tissues using the BAmSeq protocol. DNA from the original template used for each BAmSeq library was recovered after separation with streptavidin magnetic beads. The recovered DNA was then used for end-point, quantitative and droplet digital PCR (ddPCR) as well as NGS using a cancer gene panel…more »

A Survey of Molecular Heterogeneity in Hepatocellular Carcinoma

(2018), A survey of molecular heterogeneity in hepatocellular carcinoma. Hepatology Communications. doi:10.1002/hep4.1197

Understanding the heterogeneity of dysregulated pathways associated with the development of hepatocellular carcinoma (HCC) may provide prognostic and therapeutic avenues for disease management. As HCC involves a complex process of genetic and epigenetic modifications, we evaluated expression of both polyadenylated transcripts and microRNAs from HCC and liver samples derived from two cohorts of patients undergoing either partial hepatic resection or liver transplantation. Copy number variants were inferred from whole genome low‐pass sequencing data, and a set of 56 cancer‐related genes were screened using an oncology panel assay…more »

ERASE-Seq: Leveraging replicate measurements to enhance ultralow frequency variant detection in NGS data

PLoS ONE 13(4): e0195272. https://doi.org/10.1371/journal.pone.0195272

The accurate detection of ultralow allele frequency variants in DNA samples is of interest in both research and medical settings, particularly in liquid biopsies where cancer mutational status is monitored from circulating DNA. Next-generation sequencing (NGS) technologies employing molecular barcoding have shown promise but significant sensitivity and specificity improvements are still needed to detect mutations in a majority of patients before the metastatic stage. To address this we present analytical validation data for ERASE-Seq (Elimination of Recurrent Artifacts and Stochastic Errors), a method for accurate and sensitive detection of ultralow frequency DNA variants in NGS data…more »

Somatic activating mutations in MAP2K1 cause melorheostosis

Nature Communications. Volume 9, Article number: 1390 (2018) doi:10.1038/s41467-018-03720-z

Melorheostosis is a sporadic disease of uncertain etiology characterized by asymmetric bone overgrowth and functional impairment. Using whole exome sequencing, we identify somatic mosaic MAP2K1mutations in affected, but not unaffected, bone of eight unrelated patients with melorheostosis. The activating mutations (Q56P, K57E and K57N) cluster tightly in the MEK1 negative regulatory domain. Affected bone displays a mosaic pattern of increased p-ERK1/2 in osteoblast immunohistochemistry…more »

Applying Precision Medicine to Ovarian Cancer: Proof-of-Principle for a “Molecular Second Look”

Int J Gynecol Cancer. 2018 Jan 18. doi: 10.1097/IGC.0000000000001190

The objectives of this study were to assess if targeted investigation for tumor-specific mutations by ultradeep DNA sequencing of peritoneal washes of ovarian cancer patients after primary surgical debulking and chemotherapy, and clinically diagnosed as disease free, provides a more sensitive and specific method to assess actual treatment response and tailor future therapy and to compare this “molecular second look” with conventional cytology and histopathology-based findings…more »

Comparing mutational profiles between cell-free circulating tumor DNA and tumor DNA in laryngeal carcinoma patients

The incidence of laryngeal cancers is on the rise with an estimated 12,000 new cases diagnosed each year in the United States. Despite advancements in surgery, chemotherapy, radiotherapy, and targeted therapy, the prognosis for advanced laryngeal cancer patients is poor due to local invasion and metastasis. Identifying biomarkers that can predict response to therapy and potential recurrence in patients is critical to improving survival…more »

Circulating cell-free DNA mutation patterns in early and late stage colon and pancreatic cancer

DOI: http://dx.doi.org/10.1016/j.cancergen.2017.08.006

Cancer is a heterogeneous disease harboring diverse subclonal populations with different DNA mutations. We used circulating cell-free DNA (ccfDNA) to assess the mutational makeup and monitor changes during disease progression of pancreatic and colorectal cancers after surgery. A 56 cancer-associated gene panel showed that less than half of the mutations in the primary tumors were also detected in the ccfDNA. Also, additional mutations not detected in the primary tumors were found in the ccfDNA due to disease heterogeneity or metastatic spread at the time of diagnosis…more »

Novel insights into the molecular heterogeneity of hepatocellular carcinoma

bioRxiv preprint first posted online Jan. 19, 2017. doi: https://doi.org/10.1101/101766

Hepatocellular carcinoma (HCC) is influenced by numerous factors, which results in diverse genetic, epigenetic and transcriptional scenarios, thus posing obvious challenges for disease management. We scrutinized the molecular heterogeneity of HCC with a multi-omics approach in two small cohorts of resected and explanted livers. Whole-genome transcriptomics was conducted, including polyadenylated transcripts and micro (mi)-RNAs. Copy number variants (CNV) were inferred from whole genome low-pass sequencing data…more »

Brief report: MET exon 14 alterations and new resistance-mutations to tyrosine kinase inhibitors: risk of inadequate detection with current amplicon-based NGS panels

J Thorac Oncol. 2017 Aug 2. pii: S1556-0864(17)30657-3. doi: 10.1016/j.jtho.2017.07.026. [Epub ahead of print]

Targeted therapies, as tyrosine kinase inhibitors (TKI), have dramatically improved the treatment of lung adenocarcinoma and detection of activating mutations of genes like EGFR or ALK is now mandatory in clinical setting. However, additional targetable alterations are continuously described and force us to adapt our detection methods. We evaluate here the ability of 8 amplicon-based next generation sequencing (NGS) panels to detect the recently described MET exon 14 alterations or new resistance-mutations to TKI…more »

Study of preanalytic and analytic variables for clinical next-generation sequencing of circulating cell-free nucleic acid

J Mol Diagn. 2017 May 12. pii: S1525-1578(17)30027-2. doi: 10.1016/j.jmoldx.2017.03.003. [Epub ahead of print]

Detection of mutations in plasma circulating cell-free DNA (cfDNA) by next-generation sequencing (NGS) has opened up new possibilities for monitoring treatment response and disease progression in patients with solid tumors. However, implementation of cfDNA genotyping in diagnostic laboratories requires systematic assessment of preanalytical parameters and analytical performance of NGS platforms. We assessed the effects of peripheral blood collection tube and plasma separation time on cfDNA yield and integrity and performance of the Ion PGM, Proton, and MiSeq NGS platforms…. more »

Comparison of BEAMing assays and competitive approaches in the detection of main alteration of RAS in circulating DNA of non small-cell lung cancer (NSCLC) and metastatic colon cancer. Manuscript 2016

J Clin Oncol 35, no. 15_suppl – published online before print. DOI: 10.1200/JCO.2017.35.15_suppl.e23056

A number of RAS mutations confer resistance to anti-EGFR therapies routinely used in the treatment of colon cancer. The objective of this study was to evaluate the pertinence of analyzing circulating-free plasma DNA (cfDNA) as an indicator of the mutational status of a tumor, in order to use liquid biopsies instead of invasive and painful tumor biopsies during tumor progression…. more »

Genomic analysis of uterine lavage fluid detects early endometrial cancers and reveals a prevalent landscape of driver mutations in women without histopathologic evidence of cancer: a prospective cross-sectional study

PLoS Med. 2016 Dec 27;13(12):e1002206. doi: 10.1371/journal.pmed.1002206. eCollection 2016

Endometrial cancer is the most common gynecologic malignancy, and its incidence and associated mortality are increasing. Despite the immediate need to detect these cancers at an earlier stage, there is no effective screening methodology or protocol for endometrial cancer….Based on these cancer genome results, and in a prospective study, we hypothesized that the use of ultra-deep, targeted gene sequencing could detect somatic mutations…. more »

Cell-free DNA and next-generation sequencing in the service of personalized medicine for lung cancer

Oncotarget. 2016 Aug 30. DOI: 10.18632/oncotarget.11717

Personalized medicine has emerged as the future of cancer care to ensure that patients receive individualized treatment specific to their needs. In order to provide such care, molecular techniques that enable oncologists to diagnose, treat, and monitor tumors are necessary. In the field of lung cancer, cell free DNA (cfDNA) shows great potential as a less invasive liquid biopsy technique, and next-generation sequencing (NGS) is a promising tool for analysis of tumor mutations…. more »

Risk stratification of Barrett’s oesophagus using a non-endoscopic sampling method coupled with a biomarker panel: a cohort study

Lancet. 2016, Nov 10; doi: 10.1016/S2468-1253(16)30118-2.

…FFPE-extracted DNA was quantified by PCR with primers specific to ALU115 repetitive elements (appendix 1, p 1). At least 10-25 ng quantified DNA was used for library preparation with TP53 Accel-Amplicon comprehensive panel (Swift Biosciences, Ann Arbor, MI, US)…more »

Newborn screening quality assurance program for CFTR mutation detection and gene sequencing to identify Cystic Fibrosis

Journal of Inborn Errors of Metabolism & Screening. 2016, Volume 4:1-11.

All newborn screening laboratories in the United States and many worldwide screen for cystic fibrosis. Most laboratories use a second-tier genotyping assay to identify a panel of mutations in the CF transmembrane regulator (CFTR) gene… more »

Transcriptional and epigenomic landscapes of CNS and non-CNS vascular endothelial cells

eLife, 2018 Sep 6. doi:  10.7554/eLife.36187

Vascular endothelial cell (EC) function depends on appropriate organ-specific molecular and cellular specializations. To explore genomic mechanisms that control this specialization, we have analyzed and compared the transcriptome, accessible chromatin, and DNA methylome landscapes from mouse brain, liver, lung, and kidney ECs. Analysis of transcription factor (TF) gene expression and TF motifs at candidate cis-regulatory elements reveals both shared and organ-specific EC regulatory networks. In the embryo, only those ECs that are adjacent to or within the central nervous system (CNS) exhibit canonical Wnt signaling, which correlates precisely with blood-brain barrier (BBB) differentiation and Zic3expression. In the early postnatal brain, single-cell RNA-seq of purified ECs reveals (1) close relationships between veins and mitotic cells and between arteries and tip cells, (2) a division of capillary ECs into vein-like and artery-like classes, and (3) new endothelial subtype markers, including new validated tip cell markers…more.

CancerDetector: ultrasensitive and non-invasive cancer detection at the resolution of individual reads using cell-free DNA methylation sequencing data

Nucleic Acids Research, 12 June 2018, https://doi.org/10.1093/nar/gky423
The detection of tumor-derived cell-free DNA in plasma is one of the most promising directions in cancer diagnosis. The major challenge in such an approach is how to identify the tiny amount of tumor DNAs out of total cell-free DNAs in blood. Here we propose an ultrasensitive cancer detection method, termed ‘CancerDetector’, using the DNA methylation profiles of cell-free DNAs. The key of our method is to probabilistically model the joint methylation states of multiple adjacent CpG sites on an individual sequencing read, in order to exploit the pervasive nature of DNA methylation for signal amplification. Therefore, CancerDetector can sensitively identify a trace amount of tumor cfDNAs in plasma, at the level of individual reads. We evaluated CancerDetectoron the simulated data, and showed a high concordance of the predicted and true tumor fraction. Testing CancerDetector on real plasma data demonstrated its high sensitivity and specificity in detecting tumor cfDNAs…more »

Guidelines for whole genome bisulphite sequencing of intact and FFPET DNA on the Illumina HiSeq X Ten

Epigenetics & Chromatin 2018. 28 May 2018. https://doi.org/10.1186/s13072-018-0194-0
Comprehensive genome-wide DNA methylation profiling is critical to gain insights into epigenetic reprogramming during development and disease processes. Among the different genome-wide DNA methylation technologies, whole genome bisulphite sequencing (WGBS) is considered the gold standard for assaying genome-wide DNA methylation at single base resolution. However, the high sequencing cost to achieve the optimal depth of coverage limits its application in both basic and clinical research. To achieve 15× coverage of the human methylome, using WGBS, requires approximately three lanes of 100-bp-paired-end Illumina HiSeq 2500 sequencing. It is important, therefore, for advances in sequencing technologies to be developed to enable cost-effective high-coverage sequencing…more »

 

Data quality of whole genome bisulfite sequencing on Illumina platforms

Raine A, Liljedahl U, Nordlund J (2018) PLoS ONE 13(4): e0195972. https://doi.org/10.1371/journal.pone.0195972
Targeted methylation sequencing of plasma cell-free DNA (cfDNA) has a potential to expand liquid biopsies to patients with tumors without detectable oncogenic alterations, which can be potentially useful in early diagnosis. We developed a comprehensive methylation sequencing targeting 9,322 CpG sites consistently hypermethylated according to The Cancer Genome Atlas. Next, we performed a clinical validation of our method using plasma cfDNA samples from 78 patients with advanced colorectal cancer, non-small cell lung cancer (NSCLC), breast cancer or melanoma and compared results to patients’ outcomes…more »

Targeted Methylation Sequencing of Plasma Cell-free DNA for Cancer Detection and Classification

Annals of Oncology, mdy119, https://doi.org/10.1093/annonc/mdy119

The powerful HiSeq X sequencers with their patterned flowcell technology and fast turnaround times are instrumental for many large-scale genomic and epigenomic studies. However, assessment of DNA methylation by sodium bisulfite treatment results in sequencing libraries of low diversity, which may impact data quality and yield. In this report we assess the quality of WGBS data generated on the HiSeq X system in comparison with data generated on the HiSeq 2500 system and the newly released NovaSeq system. We report a systematic issue with low basecall quality scores assigned to guanines in the second read of WGBS when using certain Real Time Analysis (RTA) software versions on the HiSeq X sequencer, reminiscent of an issue that was previously reported with certain HiSeq 2500 software versions…more »

Dissecting the Functional Consequences of De Novo DNA Methylation Dynamics in Human Motor Neuron Differentiation and Physiology

Ziller et al., 2018, Cell Stem Cell 22, 1–16 April 5, 2018. https://doi.org/10.1016/j.stem.2018.02.012

The somatic DNA methylation (DNAme) landscape is established early in development but remains highly dynamic within focal regions that overlap with gene regulatory elements. The significance of these dynamic changes, particularly in the central nervous system, remains unresolved. Here, we utilize a powerful human embryonic stem cell differentiation model for the generation of motor neurons (MNs) in combination with genetic mutations in the de novo DNAme machinery. We quantitatively dissect the role of DNAme in directing somatic cell fate with high-resolution genome-wide bisulfite-, bulk-, and single-cell-RNA sequencing….more »

Setd2 deficiency impairs hematopoietic stem cell self-renewal and causes malignant transformation

Cell Research (2018) DOI:10.1038/s41422-018-0015-9

The histone H3 lysine 36 methyltransferase SETD2 is frequently mutated in various cancers, including leukemia. However, there has not been any functional model to show the contribution of SETD2 in hematopoiesis or the causal role of SETD2 mutation in tumorigenesis. In this study, using a conditional Setd2 knockout mouse model, we show that Setd2 deficiency skews hematopoietic differentiation and reduces the number of multipotent progenitors; although the number of phenotypic hematopoietic stem cells (HSCs) in Setd2-deleted mice is unchanged, functional assays, including serial BM transplantation, reveal that the self-renewal and competitiveness of HSCs are impaired….more »

Global delay in nascent strand DNA methylation

Nature Structural & Molecular Biology (2018) DOI:10.1038/s41594-018-0046-4

Cytosine methylation is widespread among organisms and essential for mammalian development. In line with early postulations of an epigenetic role in gene regulation, symmetric CpG methylation can be mitotically propagated over many generations with extraordinarily high fidelity. Here, we combine BrdU labeling and immunoprecipitation with genome-wide bisulfite sequencing to explore the inheritance of cytosine methylation onto newly replicated DNA in human cells….more »

Genome-wide tracking of dCas9-methyltransferase footprints

Nature Communications, volume 9, Article number: 597(2018); DOI:10.1038/s41467-017-02708-5

In normal mammalian development cytosine methylation is essential and is directed to specific regions of the genome. Despite notable advances through mapping its genome-wide distribution, studying the direct contribution of DNA methylation to gene and genome regulation has been limited by the lack of tools for its precise manipulation. Thus, combining the targeting capability of the CRISPR–Cas9 system with an epigenetic modifier has attracted interest in the scientific community….more »

Zika Virus Alters DNA Methylation of Neural Genes in an Organoid Model of the Developing Human Brain

Zika virus (ZIKV) infection during early pregnancy can cause microcephaly and associated defects at birth, but whether it can induce neurologic sequelae that appear later in life remains unclear. Using a model of the developing brain based on embryonic stem cell-derived brain organoids, we studied the impact of ZIKV infection on the DNA methylation pattern across the entire genome in selected neural cell types. The virus unexpectedly alters the DNA methylome of neural progenitors, astrocytes, and differentiated neurons at genes that have been implicated in the pathogenesis of a number of brain disorders, most prominently mental retardation and schizophrenia…more »

Genetic determinants and epigenetic effects of pioneer-factor occupancy

Nature Genetics (2018) DOI:10.1038/s41588-017-0034-3.

Transcription factors (TFs) direct developmental transitions by binding to target DNA sequences, influencing gene expression and establishing complex gene-regultory networks. To systematically determine the molecular components that enable or constrain TF activity, we investigated the genomic occupancy of FOXA2, GATA4 and OCT4 in several cell types. Despite their classification as pioneer factors, all three TFs exhibit cell-type-specific binding, even when supraphysiologically and ectopically expressed….more »

Enrichment methods provide a feasible approach to comprehensive and adequately powered investigations of the brain methylome

Nucleic Acids Research, Volume 45, Issue 11, 20 June 2017, Pages e97, https://doi.org/10.1093/nar/gkx143.

Methylome-wide association studies are typically performed using microarray technologies that only assay a very small fraction of the CG methylome and entirely miss two forms of methylation that are common in brain and likely of particular relevance for neuroscience and psychiatric disorders. The alternative is to use whole genome bisulfite (WGB) sequencing but this approach is not yet practically feasible with sample sizes required for adequate statistical power. …more »

ADHFE1 is a breast cancer oncogene and induces metabolic reprogramming

J Clin Invest. 2017. DOI: 10.1172/JCI93815

Metabolic reprogramming in breast tumors is linked to increases in putative oncogenic metabolites that may contribute to malignant transformation. We previously showed that accumulation of the oncometabolite, 2-hydroxyglutarate (2HG), in breast tumors was associated with MYC signaling, but not with isocitrate dehydrogenase (IDH) mutations, suggesting a distinct mechanism for increased 2HG in breast cancer….more »

Single-cell methylomes identify neuronal subtypes and regulatory elements in mammalian cortex

Science  11 Aug 2017: Vol. 357, Issue 6351, pp. 600-604. DOI: 10.1126/science.aan3351

The presence or absence of methylation on chromosomal DNA can drive or repress gene expression. Now, a comprehensive map of methylation variation in neuronal cell populations, including a between-species comparison, illustrates how epigenetic diversity plays important roles in neuronal development. Luo et al. examined how DNA methylation is both similar and different within neurons at the single-nucleus level in humans and mice. They identified 16 mouse and 21 human neuronal clusters, with greater complexity of excitatory neurons in deep brain layers than in superficial layers….more »

CancerLocator: non-invasive cancer diagnosis and tissue-of-origin prediction using methylation profiles of cell-free DNA

Genome Biology 2017 18:53. DOI:10.1186/s13059-017-1191-5

A probabilistic method, CancerLocator, exploits the diagnostic potential of cell-free DNA by determining not only the presence but also the location of tumors. CancerLocator simultaneously infers the proportions and the tissue-of-origin of tumor-derived cell-free DNA in a blood sample using genome-wide DNA methylation data. CancerLocator outperforms two established multi-class classification methods on simulations and real data, even with the low proportion of tumor-derived DNA in the cell-free DNA scenarios….more »

Cell-free DNA, inflammation, and the initiation of spontaneous term labor

American Journal of Obstetrics and Gynecology (2017), doi: 10.1016/j.ajog.2017.05.027

Hypomethylated cell-free DNA from senescent placental trophoblasts may be involved in the activation of the inflammatory cascade to initiate labor. Objective: To determine the changes in cell-free DNA concentrations, the methylation ratio, and inflammatory markers between women in labor at term vs women without labor. In this prospective cohort study, eligible participants carried a nonanomalous singleton fetus. Women with major medical comorbidity, preterm labor, progesterone use, aneuploidy, infectious disease, vaginal bleeding, abdominal trauma, or invasive procedures during the pregnancy were excluded….more »

SPlinted Ligation Adapter Tagging (SPLAT), a novel library preparation method for whole genome bisulphite sequencing

Nucleic Acids Res. 2017 Apr 7; 45(6): e36. 2016 Nov 29. doi: 10.1093/nar/gkw1110

Sodium bisulphite treatment of DNA combined with next generation sequencing (NGS) is a powerful combination for the interrogation of genome-wide DNA methylation profiles. Library preparation for whole genome bisulphite sequencing (WGBS) is challenging due to side effects of the bisulphite treatment, which leads to extensive DNA damage. Recently, a new generation of methods for bisulphite sequencing library preparation have been devised…. more »

Targeted bisulfite sequencing of the dynamic DNA methylome

Epigenetics & Chromatin 2016 9:55. Doi: 10.1186/s13072-016-0105-1.

The ability to measure DNA methylation precisely and efficiently continues to drive our understanding of this modification in development and disease. Whole genome bisulfite sequencing has the advantage of theoretically capturing all cytosines in the genome…. more »

Detecting DNA cytosine methylation using nanopore sequencing

Nat Methods. 2017 Apr;14(4):407-410. doi: 10.1038/nmeth.4184. Epub 2017 Feb 20

In nanopore sequencing devices, electrolytic current signals are sensitive to base modifications, such as 5-methylcytosine (5-mC). Here we quantified the strength of this effect for the Oxford Nanopore Technologies MinION sequencer. By using synthetically methylated DNA, we were able to train a hidden Markov model to distinguish 5-mC from unmethylated cytosine. We applied our method to sequence the methylome of human DNA, without requiring special steps for library preparation…. more »

Whole exome and target sequencing identifies MAP2K5 as novel susceptibility gene for familial non‐medullary thyroid carcinoma

International Journal of Science. 22 August 2018. DOI: 10.1002/ijc.31825.

Although the genotype‐phenotype for familial medullary thyroid carcinoma (FMTC) is well studied, only few low susceptibility risk loci were identified for familial non‐medullary thyroid carcinoma (FNMTC). The aim of this study is to screen and identify high‐penetrate genes for FNMTC. A total of 34 families with more than two first‐degree relatives diagnosed as papillary thyroid cancer without other familial syndrome were recruited. Whole exome and target gene sequencing were performed for candidate variants…more.

Diversity of DNA and RNA Viruses in Indoor Air As Assessed via Metagenomic Sequencing

Environmental Science & Technology Article ASAP. DOI: 10.1021/acs.est.7b04203

Diverse bacterial and fungal communities inhabit human-occupied buildings and circulate in indoor air; however, viral diversity in these man-made environments remains largely unknown. Here we investigated DNA and RNA viruses circulating in the air of 12 university dormitory rooms by analyzing dust accumulated over a one-year period on heating, ventilation, and air conditioning (HVAC) filters. A metagenomic sequencing approach was used to determine the identity and diversity of viral particles extracted from the HVAC filters…. more »

Detection of two non-synonymous SNPs in SLC45A2 on BTA20 as candidate causal mutations for oculocutaneous albinism in Braunvieh cattle

Genetics Selection Evolution (2017) 49:73 DOI: 10.1186/s12711-017-0349-7

Cases of albinism have been reported in several species including cattle. So far, research has identified many genes that are involved in this eye-catching phenotype. Thus, when two paternal Braunvieh half-sibs with oculocutaneous albinism were detected on a private farm, we were interested in knowing whether their phenotype was caused by an already known gene/mutation…. more »

The draft genome of the hyperthermophilic archaeon Pyrodictium delaneyi strain hulk, an iron and nitrate reducer, reveals the capacity for sulfate reduction

Stand Genomic Sci. 2017; 12: 47. ePub 2017 Aug 15. doi:  10.1186/s40793-017-0260-4

Pyrodictium delaneyi strain Hulk is a newly sequenced strain isolated from chimney samples collected from the Hulk sulfide mound on the main Endeavour Segment of the Juan de Fuca Ridge (47.9501 latitude, −129.0970 longitude, depth 2200 m) in the Northeast Pacific Ocean. The draft genome of strain Hulk shared 99.77% similarity with the complete genome of the type strain Su06T, which shares with strain Hulk the ability to reduce iron and nitrate for respiration….. more »

The 1.78-kb insertion in the 3′-untranslated region of RXFP2 does not segregate with horn status in sheep breeds with variable horn status

Genet Sel Evol. 2016 Oct 19;48(1):78

The mode of inheritance of horn status in sheep is far more complex than a superficial analysis might suggest. Observations, which were mostly based on crossbreeding experiments, indicated that the allele that results in horns is dominant in males and recessive in females, and some authors even speculated about the involvement of more than two alleles. However, all recent genome-wide association analyses point towards a very strong effect of a single autosomal locus on ovine chromosome 10…. more »

Chapter Eight – Sequencing DNA for the oxidatively modified base 8-Oxo-7,8-Dihydroguanine

Methods Enzymol. 2017;591:187-210. doi: 10.1016/bs.mie.2017.03.004. Epub 2017 Apr 7

The DNA base guanine (G) can be oxidatively modified to 8-oxo-7,8-dihydroguanine (OG). Extraction of genomic DNA followed by nuclease digestion and mass spectrometry analysis has found OG is present at background levels of ~ 1 out of 106 Gs; however, this approach cannot determine the locations for the OGs in the genome. Thus, in this methods report, we outline three different methods (A, B, and C) for sequencing OG in DNA… more »

Single-stranded DNA library preparation preferentially enriches short maternal DNA in maternal plasma

Clin Chem. 2017 May;63(5):1031-1037. doi: 10.1373/clinchem.2016.268656. Epub 2017 Mar 9

Recent studies have suggested that single-stranded DNA (ssDNA) library preparation can enrich short DNA species from the plasma of healthy individuals, cancer patients, and transplant recipients. Based on previous observations that fetal DNA molecules in the maternal plasma are shorter than maternal DNA molecules, ssDNA library preparation may potentially enrich fetal DNA and provide substantial improvement in noninvasive prenatal testing… more »

Sequencing historical specimens: successful preparation of small specimens with low amounts of degraded DNA

Mol Ecol Resour. 2017 Feb 15. doi: 10.1111/1755-0998.12660. [Epub ahead of print]

Despite advances that allow DNA sequencing of old museum specimens, sequencing small-bodied, historical specimens can be challenging and unreliable as many contain only small amounts of fragmented DNA. Dependable methods to sequence such specimens are especially critical if the specimens are unique. We attempt to sequence small-bodied (3–6 mm) historical specimens (including nomenclatural types) of beetles that have been housed, dried, in museums for 58–159 years, and for which few or no suitable replacement specimens exist… more »

Towards quantitative viromics for both double-stranded and single-stranded DNA viruses

Peer J 4:e2777. doi: 10.7717/peerj.2777

Viruses strongly influence microbial population dynamics and ecosystem functions. However, our ability to quantitatively evaluate those viral impacts is limited to the few cultivated viruses and double-stranded DNA (dsDNA) viral genomes captured in quantitative viral metagenomes (viromes). This leaves the ecology of non-dsDNA viruses nearly unknown, including single-stranded DNA (ssDNA) viruses that have been frequently observed in viromes… more »

Single-stranded DNA phages: from early molecular biology tools to recent revolutions in environmental microbiology

FEMS Microbiol Lett. 2016 Mar;363(6). pii: fnw027. doi: 10.1093/femsle/fnw027. Epub 2016 Feb 5.

Single-stranded DNA (ssDNA) phages are profoundly different from tailed phages in many aspects including the nature and size of their genome, virion size and morphology, mutation rate, involvement in horizontal gene transfer, infection dynamics and cell lysis mechanisms. Despite the importance of ssDNA phages as molecular biology tools and model systems, the environmental distribution and ecological roles of these phages have been largely unexplored… more »

GATA2 deficiency in children and adults with severe pulmonary alveolar proteinosis and hematologic disorders

BMC Pulm Med. 2015 Aug 12;15:87. doi: 10.1186/s12890-015-0083-2.

The majority of cases with severe pulmonary alveolar proteinosis (PAP) are caused by auto-antibodies against GM-CSF. A multitude of genetic and exogenous causes are responsible for few other cases… more »

Unique synteny and alternate splicing of the chitin synthases in closely related heliothine moths

Gene. 2015 Dec 10;574(1):121-39. doi: 10.1016/j.gene.2015.08.001. Epub 2015 Aug 5.

Chitin is an extracellular biopolymer that contributes to the cuticular structural matrix in arthropods. As a consequence of its rigid structure, the chitinous cuticle must be shed and replaced to accommodate growth… more »

Complete mitochondrial genome of Muricea crassa and Muricea purpurea (Anthozoa: Octocorallia) from the eastern tropical Pacific

bioRxiv. doi: http://dx.doi.org/10.1101/042945.

We sequenced the complete mitogenomes of two eastern tropical Pacific gorgonians, Muricea crassa and Muricea purpurea, using NGS technologies. The assembled mi- togenomes of M. crassa and M. purpurea were 19,586 bp and 19,358 bp in length, with a GC-content ranging from 36.0% to 36.1%, respectively… more »

Genome Sequences of Campylobacter jejuni 81-176 Variants with Enhanced Fitness Relative to the Parental Strain in the Chicken Gastrointestinal Tract

Genome Announc. 2(1):e00006-14. doi:10.1128/genomeA.00006-14.

Campylobacter jejuni is a major cause of food-borne infections in the United States due to its ability to asymptomatically colonize the gastrointestinal tracts of chickens. Using competition assays with parental C. jejuni 81-176, variants with consistently improved fitness in chicken ceca relative to the parental strain were identified and sequenced… more »