How to Make Many Copies of a Specific DNA Segment

This patent describes the Polymerase Chain Reaction (PCR), a fundamental process for making millions of copies of a specific DNA or RNA segment from a tiny sample, enabling its detection.

What it covers

This patent describes a process for detecting a specific nucleic acid sequence by repeatedly copying it. First, a sample containing nucleic acids is treated with two short DNA pieces called oligonucleotide primers, one for each strand of the target sequence, under conditions where they attach and an enzyme builds new complementary strands (Claim 1a). Next, the sample is heated to separate these newly formed strands from their templates (Claim 1b). The primers and enzyme then build new strands again using the separated strands as templates (Claim 1c). Repeating these steps (Claim 2) exponentially increases the amount of the specific nucleic acid. Finally, a labeled probe is added to bind to the amplified sequence, allowing its detection (Claim 1d, 1e). For example, this process could be used to find a specific viral DNA sequence in a patient sample.

What it doesn't cover

• —Does not cover methods that amplify DNA without using two oligonucleotide primers, one for each strand of the target sequence.
• —Does not cover methods that amplify DNA without repeating the denaturation and extension steps at least once.
• —Does not cover methods for detecting a nucleic acid sequence that do not involve adding a labeled oligonucleotide probe and determining its hybridization.
• —Does not cover amplification processes where the primers do not create extension products that can themselves serve as templates for the other primer.
• —Does not cover methods that use enzymes not listed or functionally equivalent to those described in Claim 15 for polymerization.

The clever bit

The truly novel aspect was the idea of using a pair of primers and repeated cycles of denaturation, annealing, and extension to exponentially amplify a specific DNA segment, with each newly synthesized strand serving as a template for subsequent rounds.

Why it matters

This patent covers the Polymerase Chain Reaction (PCR), a technique that revolutionized molecular biology, medicine, and forensics. It made it possible to study tiny amounts of DNA, leading to breakthroughs in genetic research, disease diagnosis, and criminal investigations. PCR became an indispensable tool across countless scientific and commercial applications.

Real-world examples

• 1.COVID-19 diagnostic tests
• 2.Forensic DNA analysis (e.g., crime scene investigation)
• 3.Paternity testing
• 4.Genetic disease screening (e.g., for sickle cell anemia as mentioned in Claim 12)
• 5.Detection of pathogenic organisms (e.g., bacteria, viruses)
• 6.Gene sequencing preparation