SUPPLEMENT ARTICLE Whole-Exome/Genome Sequencing and Genomics AUTHORS: Wayne W. Grody, MD, PhD, FCAP, FACMG,a Barry H. Thompson, MD, MS, FAAP, FACMG,b and Louanne abstract c Hudgins, MD, FAAP, FACMG As medical genetics has progressed from a descriptive entity to one aDivisions of Medical Genetics and Molecular Pathology, focused on the functional relationship between genes and clinical dis- Departments of Pathology and Laboratory Medicine, Pediatrics, and Human Genetics, UCLA School of Medicine, Los Angeles, orders, emphasis has been placed on genomics. Genomics, a subelement California; bAmerican College of Medical Genomics and Genetics, of genetics, is the study of the genome, the sum total of all the genes of Bethesda, Maryland; and cDivision of Medical Genetics, an organism. The human genome, which is contained in the 23 pairs of Department of Pediatrics, Stanford University School of Medicine/Lucile Packard Children’s Hospital, Stanford, California nuclear chromosomes and in the mitochondrial DNA of each cell, com- prises .6 billion nucleotides of genetic code. There are some 23 000 KEY WORDS cytogenetics, genetic testing, genomics, next-generation protein-coding genes, a surprisingly small fraction of the total genetic sequencing, primary care, whole-exome sequencing, whole- material, with the remainder composed of noncoding DNA, regulatory genome sequencing sequences, and introns. The Human Genome Project, launched in ABBREVIATION 1990, produced a draft of the genome in 2001 and then a finished DTC—direct-to-consumer sequence in 2003, on the 50th anniversary of the initial publication of www.pediatrics.org/cgi/doi/10.1542/peds.2013-1032E Watson and Crick’s paper on the double-helical structure of DNA. Since doi:10.1542/peds.2013-1032E then, this mass of genetic information has been translated at an ever- Accepted for publication Aug 28, 2013 increasing pace into useable knowledge applicable to clinical medi- Address correspondence to Louanne Hudgins, MD, FAAP, FACMG, cine. The recent advent of massively parallel DNA sequencing (also Division of Medical Genetics, Department of Pediatrics, Stanford known as shotgun, high-throughput, and next-generation sequencing) University School of Medicine/Lucile Packard Children’s Hospital, 300 Pasteur Dr, H315, Stanford, CA 94305-5208. E-mail: has brought whole-genome analysis into the clinic for the first time, [email protected] and most of the current applications are directed at children with PEDIATRICS (ISSN Numbers: Print, 0031-4005; Online, 1098-4275). congenital conditions that are undiagnosable by using standard ge- Copyright © 2013 by the American Academy of Pediatrics netic tests for single-gene disorders. Thus, pediatricians must be- FINANCIAL DISCLOSURE: The authors have indicated they have come familiar with this technology, what it can and cannot offer, no financial relationships relevant to this article to disclose. and its technical and ethical challenges. Here, we address the con- FUNDING: This effort was supported by grant UC7MC21713 from cepts of human genomic analysis and its clinical applicability for the Health Resources and Services Administration’s Maternal primary care providers. Pediatrics 2013;132:S211–S215 and Child Health Bureau. The Genetics in Primary Care Institute is a cooperative agreement between the American Academy of Pediatrics and the Maternal and Child Health Bureau. POTENTIAL CONFLICT OF INTEREST: The authors have indicated they have no potential conflicts of interest to disclose. PEDIATRICS Volume 132, Supplement 3, December 2013 S211 Downloaded from www.aappublications.org/news by guest on October 2, 2021 INTRODUCTION been enhanced by the development single genes, as we do in the current paper. Semantics aside, the ultimate The term “genetic testing” can encom- of molecular techniques, particularly fl goal of the field of genomics is to pass the detection and characteriza- uorescence in situ hybridization: the identify how mutations in multiple tion of any analyte that is determined useoflabeledDNAprobesthathybridize genes interact with each other and the or predicted by one’s inheritance. In the to intact chromosomes to reveal sub- environment, a goal that may not be broadest sense of the term, genetic microscopic deletions or duplications achieved for several years. testing includes, for example, mea- of a particular region of the genome. surements of cholesterol levels or Even more recently, chromosomal plasma metabolites such as amino microarrays, consisting of as many as 1 TECHNICAL ASPECTS OF DNA acids and microscopic examination of million DNA probes bound to a solid SEQUENCING support, have been used to interrogate a peripheral blood smear for sickle- For most of the .25-year history of shaped red cells or spherocytes. regions of the entire genome. This method is capable of detecting dele- clinical molecular diagnostics, and for These tests reflect the effects or signs the entire duration of the Human Ge- of genetic hyperlipidemias, errors of tions and duplications of chromosomal regions at much finer resolution than nome Project, DNA sequencing was amino acid metabolism, and hemolytic performed on semiautomated capil- afforded by standard karyotype. In fact, anemias, respectively. Such biochemical lary electrophoresis instruments by the power and diagnostic yield of genetic testing has been available for using the biochemical method known chromosomalmicroarraysaresomuch many decades. However, the most fun- as dideoxy-chain-termination, or Sanger greater that the American College of damental form of genetic testing clearly sequencing, after its inventor. This highly Medical Genetics and Genomics has is the direct analysis of the genetic accurate, although relatively slow, recommended that this test replace material itself (chromosomes, genes, method can provide the DNA sequence of karyotype analysis as the standard or the entire genome), which falls un- a targeted and limited region of 150 to first-tier test in the diagnostic evalua- der the domains of cytogenetics and 200 nucleotides in a single run of 1 to 2 tion of patients with congenital mal- molecular genetics. Biochemical ge- days. This length of DNA represents formations, nonspecific dysmorphic netics, cytogenetics, and molecular only a small fraction of the total length features, developmental delay/intellectual genetics are the 3 laboratory sub- of most genes, let alone the entire ge- disability, and autism.1 specialties recognized by the American nome, which explains why sequencing Board of Medical Genetics. Finally, a new technology called next- the first human genome took 13 years Theadventofclinicalcytogeneticsdates generation DNA sequencing has and cost approximately $3 billion. Ob- to 1959, with the discovery that children placed within reach the ultimate, most viously, an effort of that magnitude with Down syndrome have 47 chromo- fundamental, and highest-resolution could never remotely be translated into somes in their cells rather than the genetic analysis currently conceiv- a clinical test. able: the precise identification and or- normal 46, the abnormality being an One reason for the slow pace of Sanger extra copy of chromosome 21 (trisomy dering of all 6 billion nucleotides in the sequencing is that it is based on 21).Sincethattime,countlessdefectsin human genome. This technology, more daughter strand synthesis of a small chromosomal number, structure, and than any other, has opened the way to stretch of DNA that is selected by using position have been observed and as- true genomic medicine. the hybridization of specific oligonucle- sociated with various congenital (and It is important to point out some se- otide primers to just that region. The neoplastic) disorders. In a sense, chro- mantic distinctions related to the primers serve as start sites for mosomal analysis (also known as kar- evolving field of genomics. Most agree DNA polymerase to make complemen- yotype analysis) can be considered the that the term “genetics” refers to the tary strands that terminate whenever first genome-wide test because it study of single genes in isolation. Many a particular dideoxynucleotide de- includes all the nuclear genetic mate- refer to “genomics” as the study of all rivative is incorporated into the elon- rial of the cell. Such analysis is limited, genes in the genome and the inter- gating strand, and the resulting however, by the resolution of the light actions among them and their envi- sequence is deduced by measuring microscope; any deletion, duplication, ronments, as noted in the Executive the sizes of the terminated fragments insertion, or translocation that is too Summary. However, others use the on the capillary electrophoresis in- small to be observed under standard term “genomics” to describe whole- strument. In contrast, next-generation, magnification will go undetected. More exome and whole-genome sequenc- or massively parallel, sequencing recently, the field of cytogenetics has ing aimed at identifying mutations in breaks up the whole genome into .300 S212 GRODY et al Downloaded from www.aappublications.org/news by guest on October 2, 2021 SUPPLEMENT ARTICLE million small fragments, which are disorders are believed to be due to DIAGNOSIS AND DISCOVERY then universally primed for DNA poly- mutations in the coding regions, this THROUGH NEXT-GENERATION merase copying by using 4-color fluo- approach allows laboratories to focus SEQUENCING rescently labeled nucleotides and are exclusively on those regions and elim- Whether conducted by using a whole- analyzed on instruments