PCR Information & Resources for Teachers

PCR is a tool which expands the applications of other molecular biology tools by reducing the required initial nucleic acid sample size and the time required to generate much larger quantities of DNA for analysis. DNA or RNA samples which are too small to be analyzed by more traditional molecular techniques are easily amplified with PCR so that information about the source and content of the nucleic acid are generated in a short time. Sample size is often a limiting factor in nucleic acid analysis. For example, samples from crime scenes tend to be collected from a few drops of blood, a few hairs or a small bit of skin . Once these small samples are amplified by PCR, other molecular biology tools which require greater quantities of DNA can be used to analyze the DNA. This broadens the areas of application for molecular biology.

Clinical Applications

PCR is useful in establishing the unambiguous identity of a sample. A “sample” can be any source of DNA or RNA from an organism to a few cells. Samples are usually small amounts of blood or other body fluids, or a small collection of cells such as a cheek swab, a bit of bone or some hair.

Medical

• Genetic diseases can be identified pre- or post-natally
• Cancers can be identified and classified
• Identifying bacterial or viral strains for proper treatment or epidemiological studies

Food Science

• Samples can be tested for genetic engineering traits
• Bacterial contaminants and the source of contamination can be quickly identified
• Pedigrees and traits of valuable animal food stocks can be verified

Forensics

• Identification of corpses or body parts in cases of mass disaster, mass graves, or battlefield identification
• Analysis of pathogens
• PCR is a valuable tool in criminal investigation where it is used to eliminate possible suspects as well as link suspects to the scene of the crime either through a sample (blood, human hair, skin, semen) left at the scene or circumstantial links (pet hair, plant parts)

Relationships

PCR can also be used to establish relationships between samples, as in the following three examples:

1. Paternity




This method also demonstrates maternity, but maternity is less often in question.

• Identifying familial relationships in organisms such as wolves or chimpanzees
• Verifying pedigrees of valuable stock such as horses, cows and dogs
• Providing evidence for paternity in custody disputes
2. Evolution
• Archaeological analysis of samples from sites
• Developing relationships between species based on molecular phylogenetics
• Studying extinct species based on museum specimens
3. Ecology
• Validating conservation efforts by tracking rare wildlife or individual organisms
• Clearly defining unique subgroups within populations or species
• Documenting biodiversity by identifying unique species

Useful web sites

• Forensics

This is a page from the Human Genome Project and has information on forensics. This page links to a page with several very interesting cases of forensic identification many of which have current personal and political ramifications.

DNA testing

Private individuals may contract with a number of companies offering DNA testing. These companies tout their services not only for criminal investigations, but also for paternity, pre-natal testing, and identity establishment for estate claims and immigration purposes. There is also a market for veterinary uses of DNA testing including pedigree, identity and sexing birds. Search for “DNA Typing” on the internet to investigate DNA testing companies.

DNA testing raises a variety of ethical issues, specifically privacy issues which students will be faced with in their lifetimes. This site has an excellent essay on DNA testing issues and asks a number of ethical questions as well:

(http://www.accessexcellence.org/RC/AB/BA/)

1. Since non-invasive samples may be used for DNA testing (such as a toothbrush or hair), people can be tested without their knowledge or consent. Should this be legal?
2. The military now takes a blood sample from each person entering the military to aid in identifying bodies or body parts. The claim is that there will never be another “unknown soldier”. Many states are compiling criminal genetic records as well. This helps police identify repeat offenders or link a suspect in one case to another case. These records could potentially be used in a number of ways such as academic and medical studies where general information would be collected with the genetic information, but not personal identifiers, such as names or social security numbers. Other groups with profit in mind such as insurance companies or drug manufacturers might wish to access genetic information as well. Should genetic records be maintained, and if so, by whom? Who should have access to this information?
3. What limitations should be placed on pre-natal testing? Is it legitimate to test for gender, potential for developing adult onset diseases (Huntington’s, aggressive breast cancer) or fatal genetic flaws such as Lesch-Nyhan syndrome, which results in a short, painful life characterized by severe retar­dation, violence, and self-mutilation? What about non-fatal diseases or syndromes such as Down’s syndrome or galactosemia which require life long care and can generate other medical issues? What about diseases that can be treated, but at great cost, and with a probable decrease in life span such as Gaucher’s?
4. DNA testing can potentially lead to the development of a “genetic underclass”, those who cannot access DNA testing and so continue to produce children with genetic defects. What ethical considerations arise in this situation? What are the ramifications for these people economically such as lost work time, higher medical bills and special care costs? What role will insurance companies play in these issues? Will DNA testing be mandatory for health insurance, and if so, to what information will insurance companies be allowed access?
5. Many medical advances have changed previously deadly diseases into diseases that can be survived or managed for a full lifetime. Diabetes, PKU and childhood leukemia are not the absolute death sentences they were in the past. Should genetic screening for currently untreatable diseases take the place of medical advances in removing deadly diseases from the human race?

Further Investigations

Many scientific areas have applied molecular biology tools to address questions. Students who may not necessarily be interested in biology may find one of the following applications more interesting.

• Forensic anthropologists attempt to identify the remains in mass graves using a variety of methods including PCR.
• Epidemiologists studying new diseases and their sources will employ molecular biology to analyze strains and variants.
• Penguins, and many other birds, must be sexed by DNA analysis. Bird breeders, zoo keepers and veterinarians apply this technology.
• Criminal investigators use PCR to amplify DNA samples from crime scenes for further analysis.
• Plant and animal breeders wishing to verify the pedigree of their stock use PCR and other molecular biology techniques.
• Lawyers specializing in legal issues arising regarding intellectual property, privacy issues and appropriate use and interpretation of evidence in criminal cases.

Have students investigate genetic screening in their families. Who has been screened and what for? Infant testing for PKU and other metabolic diseases has been in use for about 40 years, so most of the students themselves have been screened.

Have students determine expenses associated with undiagnosed metabolic disorders including wages lost by caregivers, medical expenses (personal and Medicare), insurance costs and other costs such as special equipment or supplies required to care for the patient. Compare this cost with the cost of screening all infants. Include the cost of the test and recommended treatment for the patient.