DOCSLIB.ORG

Next-Generation Sequencing of Centenarians to Identify Genetic Variants

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Next-Generation Sequencing of Centenarians to Identify Genetic Variants Next-generation sequencing of centenarians to identify genetic variants predisposing to human longevity Dissertation In fulfillment of the requirements for the degree “Dr. rer. nat.” of the Faculty of Mathematics and Natural Sciences at Christian-Albrechts-Universität zu Kiel Submitted by Nandini Badarinarayan Research Group for Healthy Ageing Institute of Clinical Molecular Biology Kiel, July 2014 First referee: Prof. Dr. Tal Dagan Second referee: Prof. Dr. Almut Nebel th Date of oral examination: 24 October 2014 Signed: Prof. Dr. Wolfgang Duschl (Dean) “Always keep your smile. That's how I explain my long life.”- Jeanne Calment (who had the longest confirmed human lifespan in history, living to the age of 122 years, 164 days) For my family Contents I Table of Contents List of figures ...................................................................................... IV List of tables ..........................................................................................V List of abbreviations ........................................................................... VII 1 Introduction ............................................................................... 1 1.1 Longevity phenotype............................................................................... 1 1.2 Genetic epidemiology of human longevity ............................................. 4 1.3 Genetic influences on longevity .............................................................. 6 1.3.1 Findings in model organisms ..................................................................................... 6 1.3.2 Findings in humans .................................................................................................... 9 1.4 Next-generation sequencing to detect variants associated with human longevity .................................................................................... 13 1.5 Research objectives ............................................................................... 15 2 Materials ................................................................................. 18 2.1 Enzymes, kits and instruments .............................................................. 18 2.2 Online databases and software .............................................................. 19 3 Methods .................................................................................. 20 3.1 Sequencing ............................................................................................ 20 3.1.1 Study participants ..................................................................................................... 20 3.1.2 SOLiD technology ................................................................................................... 21 3.1.3 Illumina technology ................................................................................................. 22 3.2 Mapping and variant calling ................................................................. 25 3.3 Selection of variants .............................................................................. 27 3.3.1 Method 1: SNVs that may have functional impact .................................................. 27 3.3.2 Method 2: Low-frequency variants with functional impact ..................................... 28 Contents II 3.4 Genotyping and replication ................................................................... 31 3.4.1 Sequenom technology .............................................................................................. 31 3.4.2 TaqMan technology ................................................................................................. 32 3.4.3 Study population ...................................................................................................... 32 3.4.4 Statistical analysis .................................................................................................... 34 4 Results ..................................................................................... 35 4.1 Mapping, coverage and variant calling ................................................. 35 4.1.1 SOLiD technology ................................................................................................... 35 4.1.2 Illumina technology ................................................................................................. 37 4.2 Method 1: SNVs that may have a functional impact ............................ 42 4.2.1 Selection of variants ................................................................................................. 42 4.2.2 Analysis of selected SNVs ....................................................................................... 44 4.3 Method 2: Low-frequency variants with functional impact ................. 48 4.3.1 Selection of variants ................................................................................................. 48 4.3.2 Analysis of selected SNVs ....................................................................................... 54 5 Discussion ............................................................................... 60 5.1 Coverage and variant calling performance for SOLiD and Illumina sequencing data ...................................................................... 61 5.2 Challenges of next-generation sequencing ........................................... 66 5.3 Methods implemented for selection of variants .................................... 68 5.4 Potential influencing factors for association studies ............................ 72 5.4.1 Population stratification ........................................................................................... 72 5.4.2 Population-specificity .............................................................................................. 74 5.4.3 Phenotype heterogeneity .......................................................................................... 75 5.4.4 Sample sizes and statistical power ........................................................................... 75 5.5 Summary of findings ............................................................................. 77 5.5.1 Study findings .......................................................................................................... 77 5.5.2 Genetic profiles of centenarians ............................................................................... 79 5.6 Conclusion and outlook ........................................................................ 85 6 Summary ................................................................................. 89 Contents III 7 Zusammenfassung .................................................................. 90 8 References ............................................................................... 92 9 Declaration ............................................................................ 108 10 Curriculum vitae ................................................................... 109 11 Acknowledgements .............................................................. 110 12 Supplementary material ........................................................ 111 List of figures IV List of figures No. 1-1: Number of living supercentenarians ................................................................................... 3 1-2: Disease and longevity variants ............................................................................................ 4 1-3: Lifespan regulation in C. elegans........................................................................................ 8 3-1: Emulsion PCR for SOLiD™ sequencing .......................................................................... 22 3-2: Colour-space reference for SOLiD™ sequencing ............................................................ 22 3-3: Bridge amplification for Illumina sequencing. ................................................................. 23 3-4: Individuals sequenced on SOLiD and Illumina technologies for Method 1 ..................... 27 3-5: Individuals sequenced on SOLiD and Illumina technologies for Method 2 ..................... 29 4-1: Genotype concordance for SOLiD sequencing data ......................................................... 37 4-2: Genotype concordance for Illumina sequencing data ....................................................... 39 4-3: Genotype concordance for Illumina exome sequencing data ........................................... 42 4-4: Variant calling metrics for four exomes generated by CRG, Spain .................................. 43 4-5: Selection of variants for Method 1 .................................................................................... 44 4-6: Intersection of exonic variants between SOLiD and Illumina technology: ...................... 49 4-7: Selection of variants for Method 2 .................................................................................... 51 4-8: Power and sample size calculation .................................................................................... 57 5-1: Coverage plot for three samples sequenced with SOLiD technology............................... 61 5-2: Coverage plot for four samples sequenced with Illumina technology .............................. 62 5-3: Coverage plot for six exomes sequenced with Illumina technology................................. 63 12-1: SOLiD™ technology sequencing schema .................................................................... 114 12-2: Illumina technology sequencing schema .....................................................................
Load more

Recommended publications
  • Cytogenomic SNP Microarray - Fetal ARUP Test Code 2002366 Maternal Contamination Study Fetal Spec Fetal Cells
    Patient Report |FINAL Client: Example Client ABC123 Patient: Patient, Example 123 Test Drive Salt Lake City, UT 84108 DOB 2/13/1987 UNITED STATES Gender: Female Patient Identifiers: 01234567890ABCD, 012345 Physician: Doctor, Example Visit Number (FIN): 01234567890ABCD Collection Date: 00/00/0000 00:00 Cytogenomic SNP Microarray - Fetal ARUP test code 2002366 Maternal Contamination Study Fetal Spec Fetal Cells Single fetal genotype present; no maternal cells present. Fetal and maternal samples were tested using STR markers to rule out maternal cell contamination. This result has been reviewed and approved by Maternal Specimen Yes Cytogenomic SNP Microarray - Fetal Abnormal * (Ref Interval: Normal) Test Performed: Cytogenomic SNP Microarray- Fetal (ARRAY FE) Specimen Type: Direct (uncultured) villi Indication for Testing: Patient with 46,XX,t(4;13)(p16.3;q12) (Quest: EN935475D) ----------------------------------------------------------------- ----- RESULT SUMMARY Abnormal Microarray Result (Male) Unbalanced Translocation Involving Chromosomes 4 and 13 Classification: Pathogenic 4p Terminal Deletion (Wolf-Hirschhorn syndrome) Copy number change: 4p16.3p16.2 loss Size: 5.1 Mb 13q Proximal Region Deletion Copy number change: 13q11q12.12 loss Size: 6.1 Mb ----------------------------------------------------------------- ----- RESULT DESCRIPTION This analysis showed a terminal deletion (1 copy present) involving chromosome 4 within 4p16.3p16.2 and a proximal interstitial deletion (1 copy present) involving chromosome 13 within 13q11q12.12. This
    [Show full text]
  • Analysis of Trans Esnps Infers Regulatory Network Architecture
    Analysis of trans eSNPs infers regulatory network architecture Anat Kreimer Submitted in partial fulfillment of the requirements for the degree of Doctor of Philosophy in the Graduate School of Arts and Sciences COLUMBIA UNIVERSITY 2014 © 2014 Anat Kreimer All rights reserved ABSTRACT Analysis of trans eSNPs infers regulatory network architecture Anat Kreimer eSNPs are genetic variants associated with transcript expression levels. The characteristics of such variants highlight their importance and present a unique opportunity for studying gene regulation. eSNPs affect most genes and their cell type specificity can shed light on different processes that are activated in each cell. They can identify functional variants by connecting SNPs that are implicated in disease to a molecular mechanism. Examining eSNPs that are associated with distal genes can provide insights regarding the inference of regulatory networks but also presents challenges due to the high statistical burden of multiple testing. Such association studies allow: simultaneous investigation of many gene expression phenotypes without assuming any prior knowledge and identification of unknown regulators of gene expression while uncovering directionality. This thesis will focus on such distal eSNPs to map regulatory interactions between different loci and expose the architecture of the regulatory network defined by such interactions. We develop novel computational approaches and apply them to genetics-genomics data in human. We go beyond pairwise interactions to define network motifs, including regulatory modules and bi-fan structures, showing them to be prevalent in real data and exposing distinct attributes of such arrangements. We project eSNP associations onto a protein-protein interaction network to expose topological properties of eSNPs and their targets and highlight different modes of distal regulation.
    [Show full text]
  • Complement Factor H Deficiency and Endocapillary Glomerulonephritis Due to Paternal Isodisomy and a Novel Factor H Mutation
    Genes and Immunity (2011) 12, 90–99 & 2011 Macmillan Publishers Limited All rights reserved 1466-4879/11 www.nature.com/gene ORIGINAL ARTICLE Complement factor H deficiency and endocapillary glomerulonephritis due to paternal isodisomy and a novel factor H mutation L Schejbel1, IM Schmidt2, M Kirchhoff3, CB Andersen4, HV Marquart1, P Zipfel5 and P Garred1 1Department of Clinical Immunology, Laboratory of Molecular Medicine, Rigshospitalet, Copenhagen, Denmark; 2Department of Pediatrics, Rigshospitalet, Copenhagen, Denmark; 3Department of Clinical Genetics, Rigshospitalet, Copenhagen, Denmark; 4Department of Pathology, Rigshospitalet, Copenhagen, Denmark and 5Department of Infection Biology, Leibniz Institute for Natural Product Research and Infection Biology, Jena, Germany Complement factor H (CFH) is a regulator of the alternative complement activation pathway. Mutations in the CFH gene are associated with atypical hemolytic uremic syndrome, membranoproliferative glomerulonephritis type II and C3 glomerulonephritis. Here, we report a 6-month-old CFH-deficient child presenting with endocapillary glomerulonephritis rather than membranoproliferative glomerulonephritis (MPGN) or C3 glomerulonephritis. Sequence analyses showed homozygosity for a novel CFH missense mutation (Pro139Ser) associated with severely decreased CFH plasma concentration (o6%) but normal mRNA splicing and expression. The father was heterozygous carrier of the mutation, but the mother was a non-carrier. Thus, a large deletion in the maternal CFH locus or uniparental isodisomy was suspected. Polymorphic markers across chromosome 1 showed homozygosity for the paternal allele in all markers and a lack of the maternal allele in six informative markers. This combined with a comparative genomic hybridization assay demonstrated paternal isodisomy. Uniparental isodisomy increases the risk of homozygous variations in other genes on the affected chromosome.
    [Show full text]
  • Transcriptional Signature of Accessory Cells in the Lateral Line, Using the Tnk1bp1:EGFP Transgenic Zebrafish Line Behra Et Al
    Transcriptional signature of accessory cells in the lateral line, using the Tnk1bp1:EGFP transgenic zebrafish line Behra et al. Behra et al. BMC Developmental Biology 2012, 12:6 http://www.biomedcentral.com/1471-213X/12/6 (24 January 2012) Behra et al. BMC Developmental Biology 2012, 12:6 http://www.biomedcentral.com/1471-213X/12/6 RESEARCHARTICLE Open Access Transcriptional signature of accessory cells in the lateral line, using the Tnk1bp1:EGFP transgenic zebrafish line Martine Behra1,3*, Viviana E Gallardo1, John Bradsher3, Aranza Torrado3, Abdel Elkahloun1, Jennifer Idol1, Jessica Sheehy1, Seth Zonies1, Lisha Xu1, Kenna M Shaw2,5, Chie Satou4, Shin-ichi Higashijima4, Brant M Weinstein2 and Shawn M Burgess1 Abstract Background: Because of the structural and molecular similarities between the two systems, the lateral line, a fish and amphibian specific sensory organ, has been widely used in zebrafish as a model to study the development/ biology of neuroepithelia of the inner ear. Both organs have hair cells, which are the mechanoreceptor cells, and supporting cells providing other functions to the epithelium. In most vertebrates (excluding mammals), supporting cells comprise a pool of progenitors that replace damaged or dead hair cells. However, the lack of regenerative capacity in mammals is the single leading cause for acquired hearing disorders in humans. Results: In an effort to understand the regenerative process of hair cells in fish, we characterized and cloned an egfp transgenic stable fish line that trapped tnks1bp1, a highly conserved gene that has been implicated in the maintenance of telomeres’ length. We then used this Tg(tnks1bp1:EGFP) line in a FACsorting strategy combined with microarrays to identify new molecular markers for supporting cells.
    [Show full text]
  • Noelia Díaz Blanco
    Effects of environmental factors on the gonadal transcriptome of European sea bass (Dicentrarchus labrax), juvenile growth and sex ratios Noelia Díaz Blanco Ph.D. thesis 2014 Submitted in partial fulfillment of the requirements for the Ph.D. degree from the Universitat Pompeu Fabra (UPF). This work has been carried out at the Group of Biology of Reproduction (GBR), at the Department of Renewable Marine Resources of the Institute of Marine Sciences (ICM-CSIC). Thesis supervisor: Dr. Francesc Piferrer Professor d’Investigació Institut de Ciències del Mar (ICM-CSIC) i ii A mis padres A Xavi iii iv Acknowledgements This thesis has been made possible by the support of many people who in one way or another, many times unknowingly, gave me the strength to overcome this "long and winding road". First of all, I would like to thank my supervisor, Dr. Francesc Piferrer, for his patience, guidance and wise advice throughout all this Ph.D. experience. But above all, for the trust he placed on me almost seven years ago when he offered me the opportunity to be part of his team. Thanks also for teaching me how to question always everything, for sharing with me your enthusiasm for science and for giving me the opportunity of learning from you by participating in many projects, collaborations and scientific meetings. I am also thankful to my colleagues (former and present Group of Biology of Reproduction members) for your support and encouragement throughout this journey. To the “exGBRs”, thanks for helping me with my first steps into this world. Working as an undergrad with you Dr.
    [Show full text]
  • RNA Editing at Baseline and Following Endoplasmic Reticulum Stress
    RNA Editing at Baseline and Following Endoplasmic Reticulum Stress By Allison Leigh Richards A dissertation submitted in partial fulfillment of the requirements for the degree of Doctor of Philosophy (Human Genetics) in The University of Michigan 2015 Doctoral Committee: Professor Vivian G. Cheung, Chair Assistant Professor Santhi K. Ganesh Professor David Ginsburg Professor Daniel J. Klionsky Dedication To my father, mother, and Matt without whom I would never have made it ii Acknowledgements Thank you first and foremost to my dissertation mentor, Dr. Vivian Cheung. I have learned so much from you over the past several years including presentation skills such as never sighing and never saying “as you can see…” You have taught me how to think outside the box and how to create and explain my story to others. I would not be where I am today without your help and guidance. Thank you to the members of my dissertation committee (Drs. Santhi Ganesh, David Ginsburg and Daniel Klionsky) for all of your advice and support. I would also like to thank the entire Human Genetics Program, and especially JoAnn Sekiguchi and Karen Grahl, for welcoming me to the University of Michigan and making my transition so much easier. Thank you to Michael Boehnke and the Genome Science Training Program for supporting my work. A very special thank you to all of the members of the Cheung lab, past and present. Thank you to Xiaorong Wang for all of your help from the bench to advice on my career. Thank you to Zhengwei Zhu who has helped me immensely throughout my thesis even through my panic.
    [Show full text]
  • (12) Patent Application Publication (10) Pub. No.: US 2015/0072349 A1 Diamandis Et Al
    US 201500 72349A1 (19) United States (12) Patent Application Publication (10) Pub. No.: US 2015/0072349 A1 Diamandis et al. (43) Pub. Date: Mar. 12, 2015 (54) CANCER BOMARKERS AND METHODS OF (52) U.S. Cl. USE CPC. G0IN33/57484 (2013.01); G0IN 2333/705 (2013.01) (71) Applicant: University Health Network, Toronto USPC ......................................... 435/6.12: 435/7.94 (CA) (57) ABSTRACT A method of evaluating a probability a Subject has a cancer, (72) Inventors: Eleftherios P. Diamandis, Toronto diagnosing a cancer and/or monitoring cancer progression (CA); Ioannis Prassas, Toronto (CA); comprising: a. measuring an amount of a biomarker selected Shalini Makawita, Toronto (CA); from the group consisting of CUZD1 and/or LAMC2 and/or Caitlin Chrystoja, Toronto (CA); Hari the group CUZD1, LAMC2, AQP8, CELA2B, CELA3B, M. Kosanam, Maple (CA) CTRB1, CTRB2, GCG, IAPP, INS, KLK1, PNLIPRP1, PNLIPRP2, PPY, PRSS3, REG3G, SLC30A8, KLK3, NPY, (21) Appl. No.: 14/385,449 PSCA, RLN1, SLC45A3, DSP GP73, DSG2, CEACAM7, CLCA1, GPA33, LEFTY1, ZG16, IRX5, LAMP3, MFAP4, (22) PCT Fled: Mar. 15, 2013 SCGB1A1, SFTPC, TMEM100, NPY, PSCA RLN1 and/or SLC45A3 in a test sample from a subject with cancer; (86) PCT NO.: PCT/CA2O13/OOO248 wherein the cancer is pancreas cancer if CUZD1, LAMC2, S371 (c)(1), AQP8, CELA2B, CELA3B, CTRB1, CTRB2, GCG, LAPP (2) Date: Sep. 23, 2014 INS, KLK1, PNLIPRP1, PNLIPRP2, PPY, PRSS3, REG3G, SLC30A8, DSP GP73 and/or DSG2 is selected; the cancer is colon cancer if CEACAM7, CLCA1, GPA33, LEFTY 1 and/ Related U.S. Application Data or ZG16 is selected, the cancer is lung cancer if IRX5, (60) Provisional application No.
    [Show full text]
  • Mammalian Protein Expression and Characterization Tools for Next Generation Biologics
    Mammalian protein expression and characterization tools for next generation biologics next generation for tools characterization and expression protein Mammalian Doctoral Thesis in Biotechnology Mammalian protein expression and characterization tools for next generation biologics NIKLAS THALÉN ISBN TRITACBHFOU: KTH www.kth.se Stockholm, Sweden Mammalian protein expression and characterization tools for next generation biologics NIKLAS THALÉN Academic Dissertation which, with due permission of the KTH Royal Institute of Technology, is submitted for public defence for the Degree of Doctor of Philosophy on June 11th, 2021 at 10:00, F3, Lindstedsvägen 26, våningsplan 2, Sing-Sing, KTH campus, Stockholm. Doctoral Thesis in Biotechnology KTH Royal Institute of Technology Stockholm, Sweden 2021 © Niklas Thalén ISBN 978-91-7873-927-1 TRITA-CBH-FOU-2021:21 Printed by: Universitetsservice US-AB, Sweden 2021 Till Sofia Abstract Protein therapeutics are increasingly important for modern medicine. Novel recombinant proteins developed today can bind towards their target with high specificity and with low adverse effect. This has enabled the treatment of diseases that for a few years ago were deemed uncurable. Discovery of therapeutic proteins is driven through protein engineering, a field that is in constant expansion. And, through artificial construction of recombinant proteins, a large array of diseases can be defeated. The function and quality of these protein therapeutics rely on the correct folding, assembly and residue modification that occurs during their production within a living production cell host. Furthermore, producing them in large quantities are essential for accessibility of the best biopharmaceuticals available. Commonly, mammalian cells are the production host of choice when it comes to production of biopharmaceuticals.
    [Show full text]
  • Complement Factor H Related Proteins (Cfhrs)
    G Model MIMM-4208; No. of Pages 11 ARTICLE IN PRESS Molecular Immunology xxx (2013) xxx–xxx Contents lists available at SciVerse ScienceDirect Molecular Immunology jo urnal homepage: www.elsevier.com/locate/molimm Review Complement factor H related proteins (CFHRs) a,∗ a b,c b,d,e Christine Skerka , Qian Chen , Veronique Fremeaux-Bacchi , Lubka T. Roumenina a Department of Infection Biology, Leibniz Institute for Natural Product Research and Infection Biology, Jena, Germany b Centre de Recherche des Cordeliers, INSERM UMRS 872, Paris, France c Service d’Immunologie Biologique, Hôpital Européen Georges Pompidou, Paris, France d Université Paris Descartes Sorbonne Paris-Cité, Paris, France e Université Pierre et Marie Curie (UPMC-Paris-6), Paris, France a b s t r a c t a r t i c l e i n f o Factor H related proteins comprise a group of five plasma proteins: CFHR1, CFHR2, CFHR3, CFHR4 and Article history: CFHR5, and each member of this group binds to the central complement component C3b. Mutations, Received 1 May 2013 genetic deletions, duplications or rearrangements in the individual CFHR genes are associated with a Accepted 8 May 2013 Available online xxx number of diseases including atypical hemolytic uremic syndrome (aHUS), C3 glomerulopathies (C3 glomerulonephritis (C3GN), dense deposit disease (DDD) and CFHR5 nephropathy), IgA nephropathy, age related macular degeneration (AMD) and systemic lupus erythematosus (SLE). Although complement regulatory functions were attributed to most of the members of the CFHR protein family, the precise role of each CFHR protein in complement activation and the exact contribution to disease pathology is still unclear.
    [Show full text]
  • Molecular and Epigenetic Features of Melanomas and Tumor Immune
    Seremet et al. J Transl Med (2016) 14:232 DOI 10.1186/s12967-016-0990-x Journal of Translational Medicine RESEARCH Open Access Molecular and epigenetic features of melanomas and tumor immune microenvironment linked to durable remission to ipilimumab‑based immunotherapy in metastatic patients Teofila Seremet1,3*† , Alexander Koch2†, Yanina Jansen1, Max Schreuer1, Sofie Wilgenhof1, Véronique Del Marmol3, Danielle Liènard3, Kris Thielemans4, Kelly Schats5, Mark Kockx5, Wim Van Criekinge2, Pierre G. Coulie6, Tim De Meyer2, Nicolas van Baren6,7 and Bart Neyns1 Abstract Background: Ipilimumab (Ipi) improves the survival of advanced melanoma patients with an incremental long-term benefit in 10–15 % of patients. A tumor signature that correlates with this survival benefit could help optimizing indi- vidualized treatment strategies. Methods: Freshly frozen melanoma metastases were collected from patients treated with either Ipi alone (n: 7) or Ipi combined with a dendritic cell vaccine (TriMixDC-MEL) (n: 11). Samples were profiled by immunohistochemistry (IHC), whole transcriptome (RNA-seq) and methyl-DNA sequencing (MBD-seq). Results: Patients were divided in two groups according to clinical evolution: durable benefit (DB; 5 patients) and no clinical benefit (NB; 13 patients). 20 metastases were profiled by IHC and 12 were profiled by RNA- and MBD-seq. 325 genes were identified as differentially expressed between DB and NB. Many of these genes reflected a humoral and cellular immune response. MBD-seq revealed differences between DB and NB patients in the methylation of genes linked to nervous system development and neuron differentiation. DB tumors were more infiltrated by CD8+ and PD-L1+ cells than NB tumors.
    [Show full text]
  • A Genome-Wide Association Study Identifies Key Modulators of Complement Factor H Binding to Malondialdehyde-Epitopes
    A genome-wide association study identifies key modulators of complement factor H binding to malondialdehyde-epitopes Lejla Alica,b,c,1, Nikolina Papac-Milicevica,b,1,2, Darina Czamarad, Ramona B. Rudnicke, Maria Ozsvar-Kozmaa,b, Andrea Hartmanne, Michael Gurbisza, Gregor Hoermanna,f, Stefanie Haslinger-Huttera, Peter F. Zipfele,g, Christine Skerkae, Elisabeth B. Binderd, and Christoph J. Bindera,b,2 aDepartment of Laboratory Medicine, Medical University of Vienna, 1090 Vienna, Austria; bResearch Center for Molecular Medicine of the Austrian Academy of Sciences, 1090 Vienna, Austria; cDepartment of Medical Biochemistry, Faculty of Medicine, University of Sarajevo, 71000 Sarajevo, Bosnia and Herzegovina; dDepartment of Translational Research in Psychiatry, Max Planck Institute of Psychiatry, 80804 Munich, Germany; eDepartment of Infection Biology, Leibniz Institute for Natural Product Research and Infection Biology, 07745 Jena, Germany; fCentral Institute for Medical and Chemical Laboratory Diagnosis, University Hospital Innsbruck, 6020 Innsbruck, Austria; and gInstitute for Microbiology, Friedrich Schiller University, 07743 Jena, Germany Edited by Thaddeus Dryja, Harvard Medical School, Boston, MA, and approved March 17, 2020 (received for review August 12, 2019) Genetic variants within complement factor H (CFH), a major head-to-tail fashion. Moreover, its splice variant factor H-like alternative complement pathway regulator, are associated with protein 1 (FHL-1), consisting of the first seven SCRs of CFH, the development of age-related
    [Show full text]
  • Specificity of a Polyclonal Fecal Elastase ELISA for CELA3
    RESEARCH ARTICLE Specificity of a Polyclonal Fecal Elastase ELISA for CELA3 Frank Ulrich Weiss, Christoph Budde, Markus M. Lerch* University Medicine Greifswald, Department of Medicine A, Ferdinand Sauerbruch-Str., D-17475 Greifswald, Germany * [email protected] a11111 Abstract Introduction Elastase is a proteolytic pancreatic enzyme that passes through the gastrointestinal tract undergoing only limited degradation. ELISA tests to determine stool elastase concentra- tions have therefore been developed for the diagnosis of exocrine pancreatic insufficiency. OPEN ACCESS Five different isoforms of pancreatic elastase (CELA1, CELA2A, CELA2B, CELA3A, Citation: Weiss FU, Budde C, Lerch MM (2016) CELA3B) are encoded in the human genome. We have investigated three different poly- Specificity of a Polyclonal Fecal Elastase ELISA for CELA3. PLoS ONE 11(7): e0159363. doi:10.1371/ clonal antisera that are used in a commercial fecal elastase ELISA to determine their speci- journal.pone.0159363 ficity for different pancreatic elastase isoforms. Editor: Keping Xie, The University of Texas MD Anderson Cancer Center, UNITED STATES Material and Methods Received: October 9, 2015 Different polyclonal rabbit antisera against human elastase peptides (BIOSERV Diagnos- Accepted: July 2, 2016 tics GmbH, Germany) were tested by Western blot analysis of human pancreatic juice, in HEK-293 cells expressing Elastase constructs, and in the protein content of porcine pancre- Published: July 26, 2016 atin, used for treatment of exocrine pancreatic insufficiency. Copyright: © 2016 Weiss et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits Results unrestricted use, distribution, and reproduction in any In human pancreatic juice the polyclonal antisera detected proteins at the corresponding medium, provided the original author and source are size of human pancreatic elastase isoforms (~29kDa).
    [Show full text]