Genetic Testing and Screening Lesson 2

Lesson 2.1: Genetic Testing and Screening In Unit 1, we were introduced to the Smith family for the first time. Sue, the college-age daughter of James and Judy Smith, contracted bacterial meningitis and we worked towards treating her, handling the long-term effects (hearing loss) and studied how fu...

Lesson 2.1: Genetic Testing and Screening In Unit 1, we were introduced to the Smith family for the first time. Sue, the college-age daughter of James and Judy Smith, contracted bacterial meningitis and we worked towards treating her, handling the long-term effects (hearing loss) and studied how future infections by pathogens can be prevented. In Unit 2, we find out some new and exciting information for the Smith family - Judy Smith is pregnant again. Because Judy is a bit older than when her first two children were born, there are some additional risks in the form of an increased chance of inherited diseases. In Unit 2, we looked at a new category of diseases - diseases a person is born with (inherited diseases) rather than those they \"catch.\" We also look at genetic screening and testing, the value of screening and examining DNA, the importance of prenatal care, and the future of genetic technology. Current technology allows us to look past the surface of our cells and to understand their inner workings, their most important part - DNA. DNA can now be isolated from cells and \"picked apart\" to reveal disease. Genetic testing can be used to diagnose disease before a child is even born. We can test ourselves for diseases and learn the likelihood of passing them on to children. Genetic testing is the use of molecular methods (DNA sequencing with BLAST, karyotyping, etc.) to determine if someone has a genetic disorder, will develop one, or is a carrier of a genetic illness. It involves sampling a person\'s DNA and examining the chromosomes or genes for abnormalities. Genes, Chromosomes, and DNA DNA C T Chromosome G A Cell Gene Nucleus A bit of review: a chromosome is tightly coiled DNA. The human body contains 23 pairs of chromosomes: 22 pairs of autosomes and one pair of sex chromosomes. These chromosomes are inherited from your parents, and from the moment of conception (fertilization) they are your genetic code - your DNA. The chromosomes are typically only visible during cell division - the rest of the time, DNA is a jumbled mess that is invisible with a light microscope. This DNA, which forms chromosomes, holds genes. Genes are the coding sections of DNA, and their job is to provide the instructions for building proteins. Your body is composed of proteins. They are the workers of your body and are essentially responsible for every trait you have: hair color, eye color, blood type, skin color, and diseases you have. Chromosomes themselves can be the cause of disease, as can defective genes. In short, too many chromosomes - bad. Not enough chromosomes - bad. Inheriting defective DNA (bad gene) - also bad. Genetic Testing Overview By now, you probably realize that there are all kinds of things that can go wrong when a human being is created. It\'s a wonder more of us don\'t have things wrong. Because of the possibilities that exist for problems, many people have a strong desire to know whether they have diseases, could pass them to children, or if their unborn children have a disease. That is what genetic testing is all about - using DNA to help people find out what they want (and sometimes need) to know. Genetic testing is often performed by a genetic counselor. A genetic counselor is a trained professional who helps individuals and families understand and adjust to a genetic diagnosis or the possibility of having a hereditary disorder. Genetic counselors interpret family history information and educate patients and professionals about genetic diseases. As specialized counselors, these professionals help patients and families understand genetic testing options and the implications of undergoing genetic testing. In addition, genetic counselors address psychosocial and ethical issues associated with a genetic disorder and/or a genetic test result. As members of a health care team, genetic counselors serve as educators to their patients, to physicians, other health care providers, as well as to society. Genetic counseling can help a family understand the risks of having a child with a genetic disorder, the medical facts about an already diagnosed condition, and other information necessary for a person or couple to make decisions suitable to their cultural, religious, and moral genetic testing. In addition, genetic counselors address psychosocial and ethical issues associated with a genetic disorder and/or a genetic test result. As members of a health care team, genetic counselors serve as educators to their patients, to physicians, other health care providers, as well as to society. Genetic counseling can help a family understand the risks of having a child with a genetic disorder, the medical facts about an already diagnosed condition, and other information necessary for a person or couple to make decisions suitable to their cultural, religious, and moral dreamstime. beliefs. To keep things simple, they help with the testing and provide information people need to make informed choices. Types of Genetic Disorders Genetic testing reveals whether or not a DNA-based problem is present. These genetic disorders are caused by abnormalities in an individual\'s genetic material. We talked about four different types of genetic disorders: single-gene, multifactorial, chromosomal, and mitochondrial. You may remember these - if so, feel free to skip ahead! A single-gene disorder is a change or mutation in one gene. Sickle cell anemia and cystic fibrosis are good examples of these. Single-gene disorders may be classified as autosomal dominant, autosomal recessive, or sex-linked. A dominant trait is one where one copy of a gene passed to a child causes an effect in the child - like dwarfism or Huntington\'s disease. A recessive trait (sickle cell and cystic fibrosis) is one where a child must inherit the defective gene from both parents in order to express the trait. If the child only gets one copy, he or she is a carrier of the trait, but will not show it. A sex-linked trait is one that is passed on the sex chromosomes (the X or the Y). Remember that if a child inherits two x chromosomes, they\'re a girl. If a child gets an X and a Y (only dad can give a Y) the child is a boy. Sometimes, these X\'s and Y\'s contain defects. If a child inherits the defective chromosome, they are likely to express the trait. Sex-linked traits are a little confusing for some people because the rules are different for boys or girls. An x-linked trait is passed on the x chromosome. Because girls have two x chromosomes, they must inherit two defective x\'s to show an x-linked trait. If they only get 1, it\'s no big deal because they have a normal x to perform all the functions of the x chromosome. If a boy gets a defective x chromosome, though, they automatically have whatever bad trait was carried on that chromosome. This is because they only have one x chromosome, so there\'s no backup to perform x-related functions. This is why disorders like colorblindness, duchenne muscular dystrophy and hemophilia are much more common in males than females. Let\'s look at another type of inherited disorder now: multifactorial disorders. These are caused by multiple bad genes AND the environment in combination. Breast cancer is an example of this. People are more prone to breast cancer if they have certain forms of certain genes, but they are not guaranteed to inherit that disease. Their chances go up a lot if they make certain lifestyle choices like alcohol use or the use of deodorant. So, both the genes and the environment play a role in multifactorial diseases. Current research is suggesting that MOST common chronic illnesses (diabetes, alzheimer\'s, dementia, high blood pressure, etc.) are multifactorial. Mitochondrial disorders are fairly rare, and are caused by mutations in the DNA of mitochondria. If the mitochondria are defective, the body have a difficult time making ATP, which is needed to fuel all cell processes. These are ONLY passed from mother to child. Leber\'s hereditary optic neuropathy is an example of this. Chromosomal disorders involve inheriting either not enough chromosomes or extras. This happens when either a sperm or egg are made with the wrong number of chromosomes. Diseases where you inherit extra chromosomes include Down\'s syndrome. Down\'s Syndrome is also known as Trisomy 21. This is because a person with Down\'s syndrome has inherited an extra copy of chromosome 21. Tri- means three, and these people have three copies of a chromosome when they are only supposed to have two. You have probably met a person with Down\'s syndrome at some point. You know that the condition causes them to have very distinctive traits. These traits are caused by that trisomy. The extra DNA makes extra proteins, and this is what causes the unique physical features and internal problems seen in someone with Down\'s syndrome. These disorders are easily revealed with a karyotype, a picture of the chromosomes where they have been paired based on size, banding pattern, and centromere position, then arranged from biggest to smallest. Types of Genetic Screening It should be clear by now that there are all kinds of genetic disorders. Because of this, there are people out there who want or need to know if they carry these diseases, can pass them on to children, or have diseases themselves. There are several types of genetic testing and screening used to provide people with that information: Carrier screening, preimplantation genetic diagnosis, fetal screening/ prenatal diagnosis, and newborn screening. Carrier screening is a test that is typically done on adult couples who PARENTS are considering having children, and want to determine if those children could inherit any diseases. Most of the time, there is a family history of something like cystic fibrosis or Tay Sachs disease in the family that the couple wants to ensure they won\'t pass to the child. Remember that a carrier is someone who holds a bad gene, but doesn\'t show it. This process is fairly simple: a blood sample is drawn, the DNA is extracted and amplified using PCR, and the DNA undergoes testing for the diseases) they are concerned about. This may involve DNA sequencing or gel electrophoresis - sometimes both. Preimplantation Genetic Diagnosis (PGD) is a bit different. This unwroo procedure is often used by people with known autosomal dominant or sex- linked conditions that they do not want to pass on to their children. Here, eggs and sperm are harvested from prospective parents. The eggs are fertilized by the sperm in vitro (in a petri dish) and the embryos are allowed to develop to the 8-cell stage. After the embryos are that big, one single cell from each embryo is removed. The DNA is extracted from that one cell, amplified, and tested for the presence of the trait the parents do not want. Healthy embryos are selected and implanted in the mother for development. This technology has several ethical dilemmas surrounding it \- remember designer babies??? Fetal Screening/Prenatal diagnosis is performed on fetuses while Fetal Screening/Prenatal diagnosis is performed on fetuses while they are still in utero (inside mommy). Amniocentesis or chorionic villus sampling are used to extract cells from the fetus for testing. Amniocentesis involves inserting a large needle through the abdomen and into the uterus, where amniotic fluid (the fluid surrounding and protecting the baby) is removed. This fluid contains cells shed from the baby: skin cells, cells from the lining of the small intestine, or cells from the bladder. The cells in this fluid Embryotall Fags are provide the DNA needed to perform Fetus genetic testing. Typically, this procedure requires the use of ultrasound to locate the baby. It is normally performed after the baby is 14 weeks old. Chorionic villus sampling, on the other hand, can be done earlier. Here, chorionic villus cells are removed from the placenta. This is done by inserting a needle vaginally and directing that needle to the placenta. A small sample of those cells - which are identical to the cells inside the baby - are removed and used for testing. Just like with amniocentesis, ultrasound is used to locate the baby as well as the placenta so the procedure can be done safely. Both procedures carry some risk of miscarriage. Newborn screening is the testing of infants shortly after birth. A small sample of blood is taken from the baby, and DNA is isolated from it for testing purposes. Newborn screening is often used to test for inherited diseases if the parents choose not to implement measures that complete this testing while the baby is still in utero. Some don\'t want to risk miscarriage, and test their babies after birth instead. There are certain newborn screenings that are done automatically for most babies: for African Americans, sickle cell is commonly tested for; for Caucasians, cystic fibrosis may be tested for; for Ashkenazi Jews, Tay Sachs is tested for. This testing allows parents to take measures to give their children the best lives possible if a disease is present, and to start treating early. Getting Enough DNA for Testing Purposes Several times this section, we have brought up a key task that is part of genetic testing: amplifying DNA by PCR. Here, we will take some time to review that procedure. PCR stands for the polymerase chain reaction. This is a laboratory procedure that produces multiple copies of a specific DNA sequence. This can be a copy of a single gene, a large segment of DNA, or the entire genome of an individual. PCR is a three step process that usually takes place in a thermal cycler (a PCR machine - the purple thingy). Three \"ingredients\" are added to a sample of DNA so that copies can be made: Taq polymerase, DNA primers, and DNA nucleotides. The Taq polymerase and DNA nucleotides are included in a little pellet called a PCR bead, while the primer needed for the specific genes being tested for is added to it. The three ingredients are discussed in the paragraphs below as PCR is reviewed. The first step of PCR is known as denaturation. The temperature in the thermal cycler cranks up to 95 degrees C \- nearly boiling. The high temperatures break up the hydrogen bonds that hold the double-stranded DNA together. Think of a zipper being completely unzipped, with the two halves falling away from each other. Denaturation is required so that new DNA can be \"grown\". The second step of PCR is called annealing. The thermal cycler cools to 55 degrees C, and the DNA primers which were added to the DNA mixture early on with the bead are ready to do their job. Think of annealing as gluing. In this stage, the DNA primers (short sequences of DNA that target the beginning of the section of DNA being copied) bind to the section of DNA that scientists wish to copy. The primer is there so that the DNA is \"primed\" (readied) for copying. It tells Taq polymerase, described below, what section of DNA it should copy. Finally, extension occurs. The temperature here is 72 degrees C, and requires both Taq polymerase and DNA nucleotides. You may remember talking about Taq polymerase in class. This is an enzyme that originated in the bacteria of hot springs, so they are able to survive the hot temperature used in PCR. In bacteria, Taq polymerase is used to copy bacterial DNA before bacterial cells DENATURATION 95°C divide. Scientists use this polymerase to copy DNA during PCR. Taq polymerase attaches to the DNA at the site of the primer. After attaching, it flows down the DNA strand, adding complementary nucleotides to the DNA so that it becomes ANNEALING -50°C double-stranded. When Taq polymerase is done doing its job, there are two double-stranded pieces of DNA made from the original one. This three-step process repeats over and over. Each time it occurs, the amount of DNA doubles. This exponential growth of DNA allows lots of DNA to be made really really quickly. Within an hour and a half, 1 copy of DNA can be turned EXTENSION 72°C into more than 2 billion. Testing for Disease Genetic testing is not complete when DNA copies have been made. PCR makes DNA, but another process is required to use it to diagnose disease. (A) Diagnosis of disease requires healthcare professionals to look inside cells and decode the message buried in the sequence of nucleotides. The genotype, what is written in our DNA, predicts phenotype, what we see as a result of that code. Genotype is the genetic code for the traits we have - eye color, dimples, or diseases. Testing for these traits can be done with the process of gel electrophoresis. When starting from scratch, this can be a fairly complicated process. It is described in the text that follow? process begins with the isolation of DNA. Cells are taken from somewhere (blood, saliva, cheek swabbing) and the cells are lysed (blown up). The blown up cells and their contents are all mixed together, so a new procedure is used to separate the DNA from the cell waste. This is centrifugation. Centrifugation (fast spinning) separates the heavy cell components from the from other waste products (plasma, spit, etc.). At the end of the process, a small pellet of cell parts - including DNA - can be found at the bottom of the spun tube, while the supernatant (fluid on top of the pellet) is merely waste that can be discarded. To that tiny tube, a small amount of Chelex is added. Chelex forces the DNA to separate itself from the remainder of the cell waste in the tube, leaving the DNA floating in fluid. After this happens, the supernatant (which in this case contains the desired DNA) is moved to a new tube, while the pellet full of cell garbage is discarded. So far\... get cells → blow up cells - spin cells → dump waste → add Chelex - move DNA-holding supernatant to new tube Now that we have a small sample of DNA, it must be amplified with PCR. As this process is described in the previous section, I\'m not going to repeat it again here. Just know the section of DNA carrying the gene(s) of interest will be copied with PCR. DNA sequence After the section of DNA we want has been copied, we next need a way to find out if people have the \"bad\" version of it or not. It is important to do something so that the different versions of the gene can Restriction enzyme EcoR/ cuts the DNA be distinguished. This is done with restriction enzymes. Sticky end Restriction enzymes are molecular scissors that recognize specific DNA sequences and cut the nucleotide strands. This allows identification of the single nucleotide polymorphisms, or SNPs. Single nucleotide polymorphisms are tiny differences in the DNA of individuals that make them unique. They are 1- nucleotide differences in the DNA. SNPs can be the reason for disease, so being able to identify them is incredibly useful for scientists. This is one method used to detect the \"bad\" genes that cause some diseases. Let\'s try to make this simpler. Restriction enzymes are used to cut DNA. They are \"picky\" enzymes whose only job is to seek out the specific DNA they recognize and cut it. If scientists know the sequence of DNA they are testing, they can use restriction enzymes to cut it into fragments to reveal SNPs. Restriction enzymes may cut one version of the gene, but not another. They may also cause cuts in different places, creating different fragments of DNA that can be used to determine what version of the gene an individual has. TAS2R38 Bitter Taste Receptor So, we have DNA. We\'ve made copies. We\'ve cut the copies. What\'s next? We have to be PTC GENE(1002 bp) able to SEE the results. The best way to do this is CHROMOSOME 7 with gel electrophoresis. Here, an agarose gel is RIGHT PRIMER prepared and placed into a buffer solution. The gel contains wells at one end to which DNA is added. Markers (standard fragments of known lengths) are added first. Following this are the samples from the patient or patients being tested. After the DNA is placed into the wells in the agarose gel, it is charged NONIASICKILL TASTER (TT) with an electrical current that separates the fragments. This occurs because DNA has a slightly \- GGCGGGCACT \-- PCR PRODUCT (221 bp) GECGECCACL PER PRODUCl221 bp negative charge. Due to the placement of the agarose gel in the buffer, DNA is pulled based on Hell Restriction Enzyme (Recognition sequence GGCO) attractive forces to the positive end of the electrophoresis chamber. The pulling separates the DNA fragments, which are stained after the process \- GGCGGGCACT 221 b0 FRAGMENTI \_GGCGG 44 bp FRAGMENT ССАСТ- 177 bp FRAGMENT is completed, and can then be read. Part of this involves determining the size of the DNA fragments. Gel electrophorese To do this, the \"known\" bands of the markers are used as a comparison for the sizes of the other fragments. The sizes are used to figure out which gene versions (Alleles) a person has inherited. The gel results can be used to determine which version 221 bp FRAGMENT of a gene a patient has, revealing their SNPs and answering many questions about the presence of disease or the ability to pass disease on to children. The end result of this entire process is a gel that reveals the genotype of the individual being tested. It may reveal that a person is the carrier of a disease, is affected by a disease, or that he/she has nothing to worry about. Remember that genotype determines phenotype, the physical characteristics of an individual. If genotype says a person carries a disease, it means they hold the gene, but aren\'t affected themselves. If genotype says a person has two copies of a disease gene, they have a disease. Healthy Pregnancy and a Healthy Baby Recall that at the beginning of this, we revealed that Judy Smith was pregnant with an oopsie baby. Genetic testing was used to determine whether or not that baby had some sort of disease. Judy Smith underwent an amniocentesis, then chromosomal analysis with a karyotype. She and her husband did not undergo carrier screening, although that option was likely made available to them. The karyotype revealed that her baby was fine. Still, the fact that the baby was genetically normal did not mean that nothing could go wrong. If a woman is not careful during pregnancy, a baby that is perfectly healthy genetically can be born with life-long complications. Throughout the pregnancy, Judy\'s health and that of her son will be monitored. Maternal health will affect the health of the baby. The first trimester of pregnancy is especially critical, as all major body systems are formed during this time, and outside agents like alcohol, cigarette smoke, and drugs can have a drastic effect. Thankfully, Judy is smart enough to avoid these substances, but she still needs to monitor her eating habits, consume enough folate, and take her prenatal vitamins. Throughout pregnancy, chemical substances can have a nasty effect on a baby and should be avoided. Exercise and a healthy diet ensure that both mom and baby have what they need to stay healthy.