Polymerase Chain Reaction (PCR) Process and History

What is the first step in a PCR process?

Which bacterium provided the Taq polymerase enzyme used in PCR?

What major problem did the Taq polymerase enzyme solve in PCR?

Before the thermal cycler was invented, how were temperature changes managed in PCR?

Which invention led to a more efficient and streamlined PCR process?

What critical role does PCR play in disease diagnosis today?

What was the original method used by Sir Alec Jeffreys for DNA fingerprinting?

What is the purpose of restriction enzymes in DNA analysis?

Which DNA markers greatly increased the sensitivity of DNA profiling methods?

What was a drawback of the original method of DNA analysis involving restriction enzymes?

What is the basis of the 'bar-code' pattern visualized during forensic DNA analysis?

Which breakthrough greatly increased the use of DNA profiling for criminal investigation in the late 1980s and early 1990s?

PCR (Polymerase Chain Reaction)

A 3-step process: denature DNA to unwind the helix and separate strands, anneal primers to DNA strands, and elongate strands with nucleotides.
Developed into a standard tool for diagnosing and monitoring diseases, used in laboratories worldwide.
Enabled the study of individual DNA across many fields of biological science.

Advances in PCR

Isolation of Taq polymerase from Thermus aquaticus, a heat-resistant bacteria, replaced the previous polymerase that couldn't withstand high temperatures.
Development of a thermal cycler by Perkin Elmer in 1987, which minimized human interaction and streamlined the process.

DNA Fingerprinting

Sir Alec Jeffreys' work involved using restriction enzymes to detect variations in specific regions of DNA.
Restriction enzymes attach to DNA, detect restriction sequences, and hydrolyze DNA bonds, creating fragments.
Analyzing DNA fragments by DNA fingerprinting indicates which DNA samples have similar restriction sequences.

Breakthroughs in DNA Profiling

Late 1980s and early 1990s: introduction of microsatellite or short tandem repeat (STR) markers and PCR amplification with fluorescent labelling.
Increased sensitivity and use of DNA profiling methods for criminal investigations.

Original DNA Profiling Technique

Required 0.5–10μg of high-quality DNA, not always available during forensic investigations.
Involved digestion by restriction enzyme HinFI, Southern blotting hybridization, and multi-locus probing (MLP) technique.
Resulted in a 'bar-code' pattern, with individual patterns based on differences in probe sequence repeats in each DNA fragment.
Process was time-consuming, taking days to complete.

Learn about the 3-step process of PCR involving denaturation, primer annealing, and elongation, along with the key advance of using Taq polymerase. Discover how PCR revolutionized research and diagnostics in labs worldwide.