Molecular Diagnostic Technologies: Making Personalized Medicine Possible
The practice of personalized medicine – a new paradigm in which medical treatment can be tailored based on an individual’s genetic profile – is made possible by technologies that probe the structure and function of genes. Molecular diagnostics is a critical component of the expanding database linking alterations of DNA and RNA with disease, disease prognosis and therapeutic response.
Advanced Diagnostics Laboratories at National Jewish Health is equipped with advanced molecular diagnostic technology supporting clinical research, as well as thorough diagnosis of diseases with an established correlation to one’s genetic profile.
PCR and Real-Time PCR
Polymerase chain reaction (PCR) technology is the gold standard in molecular diagnostic testing. Informally known as "molecular photocopying," PCR is a technique that generates billions of copies of the nucleic acid sequences that serve as instructions for the development and functioning of living organisms. The discovery of PCR was so revolutionary that it earned biochemist Kary Banks Mullis the 1993 Nobel Prize for Chemistry.
One of the most significant innovations since its discovery is real-time PCR, in which billions of DNA or RNA sequences are simultaneously generated, detected and quantified. PCR was originally a cumbersome process, but today, real-time PCR typically takes two hours or less, starting with as little as a single molecule of genetic material.
Real-time PCR is used both for cutting-edge scientific research and for established molecular diagnostic tests. It is already widely used for:
- Diagnosis of cancer via the identification of low levels of malignant cells
- Detection of slow-growing pathogens
- Diagnosis of genetic disorders
- Rapid diagnosis of disease-causing viruses, bacteria, fungi and parasites
- Quantification of HIV, HCV, HBV and other viruses in blood (“viral load”)
Microarrays, also known as “gene chips,” consist of spots of DNA arranged in a specific order on a solid surface. Microarrays can track gene activity in response to drugs or disease stimuli as well as map gene variations. Often just a few centimeters across, a microarray can probe for hundreds of thousands of distinct DNA sequences and are capable of monitoring gene expression or sequences changes across the entire genome.
In next-generation sequencing, genomic DNA is broken into millions of random fragments that then are sequenced in parallel, generating generates an enormous volume of short sequence reads. These reads are then assembled, and compared to a reference genome to identify structural variations such as mutations or insertions/deletions. Next-generation sequencing is dramatically faster than Sanger sequencing, and in most cases, more cost efficient.
Developed in the mid-1970s, Sanger sequencing was the only sequencing technology used for the following 30 years. Sanger sequencing starts with one gene or region of the genome to produce a single lengthy sequence read. In 2001, Sanger sequencing was the method used to sequence the human genome. Today, it is most often utilized in resolving ambiguities in other molecular assay technologies such as next-generation sequencing.