As part of our mission to partner with health care providers to improve patient care, Claritas Genomics is deeply committed to creating and/or providing resources that further our community’s knowledge about genetics and genomics.
The information herein is designed to provide introductory information on basic genetics, geared towards patients.
Genes, Inheritance, and Genetic Testing: Interpretation of Genetic Test Results
You may be visiting this page because your health care provider suspects that you or your child has a genetic disorder. This can be a frightening prospect. Arming yourself with knowledge about the new terminology you’re encountering can be helpful. The goal of the educational pages on this website is to provide you with a foundation of basic genetic terminology and concepts. If you have questions, contact your genetics health care provider for a more in-depth conversation.
One of the first things to know is that there is no single hard and fast rule that defines all genetic disorders except that they are caused by changes in a person’s DNA. Here are some things to know about genetic disorders:
- There is no common age at which genetic disorders develop.
Some genetic disorders develop during adulthood. One example is Huntington disease. However, many disorders that are genetic develop early in life. In fact, disorders that develop in childhood are often genetic in basis. One example of this is sickle cell anemia.
- Inheritance is key feature of a genetic disorder.
Usually, a genetic disorder is inherited from a parent. For example, Phenylketonuria, or PKU, is often inherited from carrier parents. There are genetic disorders, such as Trisomy 13, that are not inherited.
- Usually in a genetic disorder, more than one organ system is affected.
Even this is not a hard and fast rule. For example, Marfan syndrome, can affect many organs, including the heart, the eyes, and the skeletal system. In contrast, retinoblastoma usually only affects the eyes.
Many people new to the world of genetic disorders want to know how genetic disorders happen. Genetic disorders are caused by changes in a person’s DNA. DNA is the set of molecules in every cell that encode the instructions for how cells create proteins and other molecules that are the basis for life. DNA is passed from parent to child and is responsible for traits that are inherited. Genetics can therefore be considered the study of heredity, and the traits and properties that are encoded by an individual’s DNA.
There are many types of changes in DNA that cause genetic disorders- from as small as a single change in a gene, to as large as a missing or extra chromosome. All humans have variations in their DNA that cause one person to be different from another person; even identical twins have some differences in their DNA. But some variants are mutations, meaning that researchers have determined that the variation isn’t considered to be “within normal limits.”
Genetic testing helps to identify a mutation that causes the disorder and so allows the health care provider, the patient, and the family learn more about the condition and what to expect. The health care team and the family can work together to discuss next steps such as treatment options, or to discuss which other specialists need to be seen. Also, the patient’s family members can be counseled about their risk of inheriting the disease and they may choose to have genetic testing performed to determine whether they carry the same genetic change. Furthermore, this knowledge of the genetic disease helps families and patients to find other people who are managing the same disease or researchers who are studying it.
Claritas Genomics tests samples from patients who are suspected by their health care provider to have a genetic disorder. The health care provider orders a specific test, obtains the sample from the patient, and sends the sample to Claritas. At Claritas, we perform the testing that has been requested and return the results to the provider. It is our goal to support the health care provider from the beginning of the ordering process in which we help determine what test would be most appropriate to order, to the delivery of results and beyond. We aim to support health care providers and patients so that they may better navigate the complex world of genetic testing.
From gene to genome
A genetic disorder is an illness that is caused by variation in the DNA sequence of one or more genes. A patient with a genetic disorder often has signs and symptoms from birth, or develops them at an early age. Families bring their child to see a genetic specialist when the pediatrician recognizes that the symptoms are not due to a virus or a bacteria or other common childhood disease.
A gene is a single unit within a DNA sequence that instructs cells to make a protein. Much like a long length of yarn is rolled up on itself into a ball, genes are packaged into structures called chromosomes, which reside within the nucleus of nearly every cell in the body. Humans have a total of 46 chromosomes: 1 pair of sex chromosomes (Chromosomes X or Y) and 22 pairs of autosomes (non-sex chromosomes). Both males and females inherit one set of autosomes (Chromosomes 1-22) from their mother and one set from their father. This means that every person carries two copies of each autosomal gene, one from each parent. The 23rd pair of chromosomes are the sex chromosomes. With few exceptions, females inherit an X chromosome from each parent, while males inherit an X chromosome from their mother and a Y chromosome from their father.
The entirety of the genetic material contained within all 23 pairs of chromosomes is called the genome. The genome consists of over 3 billion DNA base pairs. Base pairs are the building blocks of the DNA code, Adenine (A), Guanine (G), Cytosine (C) and Thiamine (T). Of the 3 billion base pairs that make up the human genome, only a small portion (about 1-2%) actually encodes the proteins that perform the body’s functions. This small portion of the genome is referred to as the exome; the exome represents only the coding regions – known as exons – of the approximately 21,000 genes in the human genome. Genetic testing usually targets either the exome (using a process called whole exome sequencing) or individual genes within the exome.
Some genetic disorders are “monogenic”, meaning that only one gene is involved. One such monogenic disease is cystic fibrosis, a condition caused by mutations in the cystic fibrosis transmembrane conductance regulatory (called CFTR). Other genetic diseases are multigenic or polygenic, in which more than just one gene is involved. Examples of multigenic or polygenic diseases are cleft palate and heart disease.
Claritas Genomics offers a variety of genetic tests performed on different technology platforms, including next generation sequencing-based tests, deletion/ duplication assays, and microarray. To learn more about next generation sequencing (NGS) and the tests that utilize NGS technologies, click here.
While most patients inherit a disease-causing mutation from one or both parents, sometimes a mutation can be de novo, meaning that it arises for the first time in a particular individual. Testing family members can determine whether a mutation is inherited or de novo, and identify other members of the family who are at risk for a disorder.
Inherited genetic disorders are transmitted from a parent to a child at the time of conception. Identifying the inheritance pattern that a genetic disorder follows is helpful in determining the chance that the disorder will occur again in a family or whether others are at risk of having the disorder. The easiest way to determine the inheritance pattern of a disorder within a particular family is to draw a pedigree, or a family tree depiction that shows the inheritance pattern of a family trait or disorder.
Some of the basic inheritance patterns include:
- Autosomal dominant: ‘Autosomal’ refers to the mutation residing in a gene on one of the non-sex chromosomes, and ‘dominant’ means that a mutation on only one copy of the gene is sufficient to cause the disease. Autosomal dominant disorders are passed directly from parent to child, and a pedigree will often show multiple affected generations; for example, grandparents, parents, and children may be affected. In this pattern of inheritance, equal numbers of males and females would be affected; we don’t expect to see only males or only females affected. Also in this model, 50% (one-half) of an affected person’s children would inherit the mutation.
- Autosomal recessive: In order to cause an autosomal recessive disorder, both copies of an autosomal gene must have a mutation. A patient with an autosomal recessive disorder generally has inherited one disease-causing mutation from each parent. A person with one mutation is called a disease “carrier”. They typically do not show any signs of the disorder. In a disorder that has an autosomal recessive inheritance pattern, not every generation will be affected in the family tree, but there may be multiple affected people within one generation, such as siblings and cousins. Similarly to autosomal dominant disorders as described above, males and females are affected in equal numbers. Children of two carrier parents have a 25% (or a ‘one out of four’) chance of inheriting both disease-causing variants and thus showing signs of the disorder.
- X-linked dominant: ‘X-linked’ refers to a disease-causing mutation residing on the X chromosome, which is one of the chromosomes that determines the sex of an individual. In the X-linked dominant inheritance pattern, a single mutation in an X-chromosome gene is sufficient to cause the disorder. A father with an X-linked dominant disorder will transmit the affected X chromosome to all (100%) of his daughters, but because he passes only his Y chromosome to his sons, none of his sons will be affected. A mother with an X-linked dominant disorder will have a 50% chance of transmitting the affected X-chromosome, so about half of her children, whether male or female, will be affected.
- X-linked recessive: A mutation on all available copies of an X-chromosome gene is necessary to cause X-linked recessive disease. Since males only have one X chromosome, one mutation is sufficient for a male to be affected. Females, on the other hand, would require a mutation on both copies of an X-linked gene in order to be affected. Females that carry only one copy of a mutation associated with X-linked recessive disease are said to be carriers, and generally do not exhibit any symptoms. A female carrier will transmit the mutation – and the disorder – to all of her male children, while on average, 50% (or half) of her female children will be carriers and the other 50% (the other half) will not inherit the mutation.
- Y-linked: “Y-linked” refers to a genetic disorder that is caused by a mutation on the Y chromosome. Disorders that are Y-linked appear in males because males have a Y chromosome while females do not. A male with a mutation on the Y chromosome will pass the mutation to all (100%) of his male children and none (0%) of his female children.
- Mitochondrial: Mitochondria are organelles within our cells that produce the energy that cells need to function. Mitochondria are only inherited from a person’s mother, so mitochondrial inheritance is also sometimes called maternal inheritance. Male children and female children both inherit mitochondrial DNA from their mother, but only females pass mitochondrial DNA to their children.
Sometimes, a pedigree will not exhibit a classic inheritance pattern, even if a mutation is known to occur within the family. Some genetic disorders can have different degrees of penetrance, meaning that not everyone who has a mutation associated with the disorder will actually exhibit signs of that disorder. Genetic disorders can also have variable expressivity, meaning that individuals with the same mutation may express different combinations of symptoms, or first show signs of the disorder at varying ages. Finally, it is possible that a mutation appears in some, but not all, cells of a person’s body. This genetic mosaicism can alter expression of the disorder as well as inheritance, and may make it more difficult for a mutation to be detected by genetic testing. Also, it’s important to note that some genetic disorders involve more than one gene (“multigenic” or “polygenic”), or may be influenced by lifestyle or environmental factors (“multi-factorial”). These conditions, such as coronary artery disease and high blood pressure, are unlikely to follow one of the inheritance patterns listed above.
Click the links for more information about DNA sequencing and about interpreting genetic test results. If you have more questions, contact your health care provider or find a genetic counselor at www.NSGC.org.
What is Sanger sequencing?
First described by Frederick Sanger and colleagues in 1977, Sanger sequencing was the most widely used DNA sequencing method for over 25 years. The basic building block of DNA is the nucleotide. The nucleotide consists of one of four nitrogenous bases, Adenine (A), Cytosine (C), Guanine (G) or Thiamine (T), a deoxyribose sugar and a phosphate group. In a human cell, a strand of DNA is built by special enzymes called DNA polymerases that bind the phosphate group from one nucleotide to a carbon from the sugar of another nucleotide. DNA is double-stranded and forms base pairs, A with T and C with G. DNA polymerase “reads” one strand, called the template strand, and uses it to build the complementary strand.
DNA can also be synthesized in a test tube by placing the four different types of nucleotides into a tube with the DNA to be sequenced and adding the polymerase enzyme. DNA polymerase will continue to bind nucleotides together according to the template strand until there are no free-floating nucleotides left in the solution.
The Sanger sequencing method is based on the discovery that certain alterations to the phosphate group on a nucleotide will cause the polymerase to stop adding bases and fall off the template. The modified phosphate groups on the different nucleotides (A, T, C, and G) can be labeled for detection (previously radioactive labels were used, now fluorescent labels are used).
Once labeled, scientists can determine the order in which the bases are added (they can read the DNA “sentence”) because they could see where the fluorescent labels were added. In one tube, all the As fluoresced and in another tube, all the Cs fluoresced. By separating the DNA fragments by length and putting the results right next to each other, scientists can easily read which base came next, thereby deducing the DNA sequence.
Sanger sequencing is a very accurate method of DNA sequencing. However, because it is slow and expensive, it is not a feasible method for sequencing large amounts of DNA in a timely manner. Most clinical DNA sequencing today is accomplished by next generation sequencing techniques.
What is Next-Generation Sequencing?
Next-generation sequencing (NGS), sometimes called massively parallel DNA sequencing, produces thousands to millions of DNA sequencing reactions at the same time. A single gene or a large number of genes can be sequenced quickly and in a cost-effective manner, making NGS an effective high-throughput method for clinical DNA testing. In NGS, a patient’s DNA is sheared into many small fragments, and each of these fragments is sequenced many times over. The millions of sequencing reads are then mapped to the reference sequence, which serves as a standard, or a control, because the sequence is already known. Variations in a patient’s DNA are detected when enough of the sequencing reads from the patient’s DNA differs from the reference sequence.
NGS is a very accurate way to detect single nucleotide substitutions (patient has a C at a particular position, while the reference sequence has a G, for example). However, NGS is not as accurate at detecting small insertions and deletions, or correctly reading homopolymers. A homopolymer is a DNA sequence in which one base is repeated several times in a row; for example TTTTTTTTTT. Because of this limitation, most laboratories that test patient samples do a second test to confirm that the results are accurate.
NGS is also not able to reliably detect large insertions of DNA (>20 base pairs), large deletions of DNA (>50 base pairs), or copy number variants (CNVs) and other structural variants. No single test platform is perfect and it can be necessary to combine platforms for the best results.
In addition to offering NGS for many genes, Claritas also offers separate tests to detect missing or extra pieces of genes. These are known as deletion/duplication assays and are only performed for genes in which large deletions and/or duplications are known to be associated with disease. In addition, the Claritas Genomics offers the ClariView Array, a chromosome microarray test that assesses for copy number variants across the genome.
Interpretation of Genetic Results
What is a variant?
A variant is any difference in an individual’s DNA sequence when compared to a reference sequence. Variants can range from changes involving just one or a few base pairs (a substitution of one single DNA base letter for another, for example), to alterations involving long stretches of DNA (deletion of an entire portion of a gene, for instance). Only a very small amount of DNA varies between any two humans- about 0.1%.
But as the human genome is huge – 3 billion base pairs – even a 0.1% variation means that there are 3 million sites of variation between any two individuals. The vast majority of these variants are said to be silent, meaning that they do not have any biological relevance. Many variants are involved in expression of unique traits between individuals, and are responsible for differences in appearance or other heritable traits. Very few variants, perhaps only a small handful in any one individual, are associated with genetic disease.
Many variants are common in the general population – a variant that is present in more than 1% of the population is called a polymorphism. Mutations, on the other hand, are defined as variants that are present in less than 1% of the population. Generally speaking, the more rare a variant, the more likely it is to be disease-causing. Variants may exhibit different frequencies in different ancestral populations. A disease-causing variant that is rare in most of the world’s populations but more common in a particular ancestral population is called a founder mutation.
How are variants analyzed?
With the vast degree of variation in the human genome, analyzing DNA sequencing data can be a daunting task. Claritas Genomics utilizes a combination of state-of-the-art software, well-vetted algorithms, and clinical expertise to sort through millions of DNA variants in order to isolate particular disease-causing variants that are likely to be responsible for a patient’s clinical symptoms. Sometimes, an identified variant will have been previously associated with disease in the medical literature and are known to be pathogenic. For variants that haven’t been reported in the medical literature, several parameters can serve as clues as to whether a variant is pathogenic. Examples of these parameters include a variant’s gene location, population frequency, and effect on protein function.
Our ability to assess and catalog DNA variants will improve over time. As more and more patients have their DNA sequenced, databases that link a patient’s genomic data (genotype) with clinical signs and symptoms (phenotype) will become larger and more refined. This underscores the critical importance of continued research to further ascertain genotype-phenotype correlations.
What are possible results?
For a whole exome, gene panel, or single gene DNA sequencing test, there are five ways in which Claritas classifies and reports variants:
- Pathogenic variant: Based upon previous reports in the scientific literature and information in gene variant databases, this variant is a recognized cause of some or all of the patient’s clinical signs and symptoms (the patient’s phenotype).
- Likely pathogenic variant: This is a novel (previously unreported) variant, but based on its gene location and the predicted effect on protein function, it is likely to be the cause of some or all of the patient’s phenotype.
- Benign variant: Based upon previous reports in the scientific literature and information in gene variant databases, this variant is recognized as a benign (not harmful), neutral variant and is not responsible for the patient’s phenotype.
- Likely benign variant: This is a novel variant, but based on its gene location and predicted lack of effect on protein function, it is likely to be benign and not responsible for the patient’s phenotype.
- Variant of uncertain significance (VUS): Based on current evidence, it cannot be determined if this variant is pathogenic or benign. Some VUSes have been previously reported in the scientific literature but the evidence associating them with disease may be conflicting. Other VUSes are novel variants whose effect on protein function cannot be predicted. Claritas makes a concerted effort to further classify VUSes as new evidence emerges, and will notify the ordering health care provider if new information becomes available about a VUS that changes its classification. Testing parental samples can sometimes be useful to help assess the clinical significance of a VUS. If a VUS is not inherited from either parent, then this VUS is generally thought to be pathogenic. But many genetic specialists argue that this alone isn’t enough to state that the VUS causes the disorder. Likewise, if the parent carries the same VUS but does not have the same genetic disorder, then many would say that the VUS must be benign. However, due to possible reduced penetrance or variable expressivity, others argue that this isn’t enough to say that the VUS is benign. In some cases, Claritas Genomics offers free parental testing to help with the interpretation of VUSes.