Disease Gene Identification: Methods and Protocols PDF
Preview Disease Gene Identification: Methods and Protocols
M M B ™ ethods in olecular iology Series Editor John M. Walker School of Life Sciences University of Hertfordshire Hatfield, Hertfordshire, AL10 9AB, UK For other titles published in this series, go to www.springer.com/series/7651 wwwwwwwwwwww Disease Gene Identification Methods and Protocols Edited by Johanna K. DiStefano Diabetes, Cardiovascular, and Metabolic Diseases Division, Translational Genomics Research Institute, Phoenix, AZ, USA Editor Johanna K. DiStefano, Ph.D. Diabetes, Cardiovascular, and Metabolic Diseases Division Translational Genomics Research Institute Phoenix, AZ USA [email protected] ISSN 1064-3745 e-ISSN 1940-6029 ISBN 978-1-61737-953-6 e-ISBN 978-1-61737-954-3 DOI 10.1007/978-1-61737-954-3 Springer New York Dordrecht Heidelberg London © Springer Science+Business Media, LLC 2011 All rights reserved. This work may not be translated or copied in whole or in part without the written permission of the publisher (Humana Press, c/o Springer Science+Business Media, LLC, 233 Spring Street, New York, NY 10013, USA), except for brief excerpts in connection with reviews or scholarly analysis. Use in connection with any form of information storage and retrieval, electronic adaptation, computer software, or by similar or dissimilar methodology now known or hereafter developed is forbidden. The use in this publication of trade names, trademarks, service marks, and similar terms, even if they are not identified as such, is not to be taken as an expression of opinion as to whether or not they are subject to proprietary rights. Printed on acid-free paper Humana Press is part of Springer Science+Business Media (www.springer.com) Dedication This book is dedicated in memory of Stephanie Savage, a colleague and friend who touched the hearts of many and devoted her work to improving the lives of others. v wwwwwwwwwwww Preface Completion of the Human Genome Project (HGP) has provided us with a greatly enhanced understanding of human genetics, including a greater appreciation of how DNA shapes species development and evolution, biology, and disease susceptibility. The HGP has also affected the development and/or maturation of research disciplines such as genome annotation, knowledge of genome evolution and segmental duplication, and comparative genomics, among others. Yet, perhaps the greatest impact of the HGP has been on the manner in which researchers investigate the causes of complex human dis- eases. Completion of the HGP gave rise to the development of efforts to characterize genetic variation in the human genome, which has lead directly to the application of whole genome association studies to identify common alleles which contribute to complex disease risk. Efforts to identify genetic mutations underlying highly penetrant diseases have been widely successful due to the facts that (1) a single mutation is enough to cause the disease (i.e., monogenic) and (2) the mutation is inherited in a simple manner between genera- tions in affected families. To date, more than a thousand genes for such disorders have been identified. These monogenic diseases are rare compared to diseases such as type 2 diabetes mellitus, cardiovascular disease, and cancer, which have complex etiologies. Unlike monogenic diseases, which arise due to a single genetic aberration, complex dis- eases result from a complicated interaction of multiple genetic and environmental deter- minants, none of which are amenable to identification and characterization using the traditional approaches to monogenic disease gene discovery. Recent efforts to characterize genetic variation in the human genome, coupled with the rapidly developing field of genomics, have lead directly to the development of new and innovative approaches to the identification of genes contributing to complex human diseases. This volume is written to provide up-to-date molecular methodologies used in the process of identifying a disease gene, from the initial stage of study design to the next stage of preliminary locus identifi- cation, and ending with stages involved in target characterization and validation. The need for such a book derives from the intellectual revolution in biomedical science and the realization that the molecular determinants of human diseases are rapidly becoming iden- tifiable through well-planned, technologically advanced approaches. Although the research literature is replete with descriptions of technical procedures, there is typically a dearth of extensive practical detail in these publications, particularly in terms of modifications developed from personal experience, as well as discussions of potential problems that may be encountered throughout the protocol and ways to avoid them. The structure of this volume is unique in that it aims to address these deficiencies. The chapters contained within have been contributed by experts in their fields, who have kindly accepted the invitation to compile the protocols within this volume and share with us their expertise, experience, and results. This text is written at a level accessible to graduate students, postdoctoral researchers, and bench scientists in the fields of molecular genetics and molecular biology. The primary aim of vii viii Preface this volume is to present detailed laboratory procedures in an easy-to-follow format that can be carried out with success by competent investigators lacking previous exposure to a specific research method. The book’s main focus is on the application of molecular approaches to disease gene identification, but overviews and case studies are also presented. The volume begins with three introductory chapters which provide an overview of disease gene identification strategies and a description of study sample selection and suc- cessful experimental design. The next section of the text contains chapters presenting methods for identifying potential susceptibility loci, including practical procedures for high-throughput SNP genotyping using whole genome arrays, medium-throughput SNP genotyping using mass spectrometry, and low-throughput, targeted SNP genotyping approaches commonly used in fine-mapping and candidate gene investigations. The sec- tion ends with a chapter on bar-coded, multiplexed sequencing of targeted DNA regions to pinpoint specific allelic variants which contribute to disease risk. The volume follows with a section on current applications in human genomics, which provide tools for target validation and functional assessment. These protocols are typically associated with the steps of disease gene identification pursuant to initial locus discovery, such as those pertaining to functional characterization of susceptibility alleles and loci. Examples of such approaches include global mRNA expression profiling using mainstream platforms, the newly emerging field of microRNA profiling, and allelic expression profil- ing, as well as quantitative PCR and RNA mapping methods. Chapters on comparative genomic hybridization, which is a molecular-cytogenetic method to detect copy number changes, and high content analysis are also included. Finally, we end with four discursive chapters, to provide examples of disease gene identification and application. The first chapter in this section is related to bioinformatics approaches to the elucidation of gene identity and function, with a particular focus on an integrative systems biology approach. The second chapter in this section illustrates the entire process of disease gene identification with a real-life case study, and the concluding chapter presents an RNA-interference approach to functional pharmacogenomics and an application of molecular diagnostics to the treatment of b-thalassemia. Without the support and contributions of many individuals, completion of this vol- ume would not have been possible. In particular, I thank Dr. John Walker, the editor of this series at Humana Press, who ensured the smooth and effortless completion of this project from the very start. I also express gratitude to the authors, all of whom contri- buted outstanding chapters, emanating from years of experience in the field. It was a plea- sure working with this expert team of scientists, and I would gladly do so again at a moment’s notice. It is my hope that this volume leads to the identification and character- ization of many more disease-related genes, which may someday pave the way toward more accurate and improved methods for disease diagnosis, as well as novel and effective strategies for disease treatment and prevention. Phoenix, AZ, USA Johanna K. DiStefano Contents Preface............................................................. vii Contributors......................................................... xi Part I IntroductIon 1 Technological Issues and Experimental Design of Gene Association Studies ......................................... 3 Johanna K. DiStefano and Darin M. Taverna 2 Statistical Issues in Gene Association Studies............................. 17 Richard M. Watanabe 3 Identification of Causal Sequence Variants of Disease in the Next Generation Sequencing Era ................................ 37 Christopher B. Kingsley Part II Methods for Gene IdentIfIcatIon 4 Microarray-Based Genome-Wide Association Studies Using Pooled DNA ......................................... 49 Szabolcs Szelinger, John V. Pearson, and David W. Craig 5 Medium-Throughput SNP Genotyping Using Mass Spectrometry: Multiplex SNP Genotyping Using the iPLEX® Gold Assay.................. 61 Meredith P. Millis 6 Targeted SNP Genotyping Using the TaqMan® Assay ..................... 77 Dorit Schleinitz, Johanna K. DiStefano, and Peter Kovacs 7 Bar-Coded, Multiplexed Sequencing of Targeted DNA Regions Using the Illumina Genome Analyzer ........................... 89 Szabolcs Szelinger, Ahmet Kurdoglu, and David W. Craig Part III f unctIonal characterIzatIon of suscePtIbIlIty alleles and locI 8 Site-Directed Mutagenesis .......................................... 107 Patricia E. Carrigan, Petek Ballar, and Sukru Tuzmen 9 Gene Expression Profiling of Tissues and Cell Lines: A Dual-Color Microarray Method .................................... 125 Sonsoles Shack 10 Methods for MicroRNA Microarray Profiling............................ 145 Aarati R. Ranade and Glen J. Weiss 11 Allelic Expression Profiling to Dissect Genome-Wide Association Study Signals ........................................... 153 Jonathan D. Gruber ix x Contents 12 Quantitative Polymerase Chain Reaction Using the Comparative C Method......................................... 171 q Kimberly Yeatts 13 Genomic Analysis by Oligonucleotide Array Comparative Genomic Hybridization Utilizing Formalin-Fixed, Paraffin-Embedded Tissues.......................................... 185 Stephanie J. Savage and Galen Hostetter 14 RNA Mapping Protocols: Northern Blot and Amplification of cDNA Ends ..................................... 199 Maria Lucrecia Alvarez and Mahtab Nourbakhsh 15 High-Content RNA Interference Assay: Analysis of Tau Hyperphosphorylation as a Generic Paradigm ...................... 221 RiLee H. Robeson and Travis Dunckley Part IV alternatIVe aPProaches 16 Integrative Systems Biology Approaches to Identify and Prioritize Disease and Drug Candidate Genes......................... 241 Vivek Kaimal, Divya Sardana, Eric E. Bardes, Ranga Chandra Gudivada, Jing Chen, and Anil G. Jegga 17 Identification of a Common Variant Affecting Human Episodic Memory Performance Using a Pooled Genome-Wide Association Approach: A Case Study of Disease Gene Identification .................... 261 Traci L. Pawlowski and Matthew J. Huentelman 18 RNAi-Based Functional Pharmacogenomics ............................. 271 Sukru Tuzmen, Pinar Tuzmen, Shilpi Arora, Spyro Mousses, and David Azorsa 19 Genetic Predisposition to b-Thalassemia and Sickle Cell Anemia in Turkey: A Molecular Diagnostic Approach...................... 291 A. Nazli Basak and Sukru Tuzmen Erratum. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . E1 Index. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 309
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