PCR & Sequencing

Polymerase chain reaction (PCR)

A commonly used method to make millions of genetic copies of specific DNA samples. This allows scientists to amplify small samples of DNA and make them large enough to analyze.

DNA Sequencing

A process used to determine the sequence of nucleotides in a DNA sample. There are two types of sequencing, Sanger Sequencing and Next-generation Sequencing. Sequencing breaks the genome of a DNA into smaller segments, sequencing those segments and assembling the sequences into one long strand.

Sanger sequencing copies the target DNA and make fragments of different lengths. Fluorescent nucleotides mark the ends of the fragments and allow the sequence to be determined.

Next-generation sequencing are new methods for sequencing. This approach increases the speed and is cost-effective.

How does PCR work?

To amplify the sample of DNA, the sample is first heated so the DNA separates into two single stranded DNA. An enzyme called Taq polymerase builds two new strands of DNA. The two original strands of DNA act as templates for the enzyme. Each of the strands are then used to continually create two new copies. The separation and synthesizing process repeats as many as 30 times. This makes over one billion copies of the original DNA segments.

The PCR process uses an automated thermocycler that is programmed to alter the temperature of the reaction every few minutes to allow for DNA separation and synthesis.

How does sequencing work

Sanger Sequencing

Sanger Sequencing

Fragments of DNA are aligned by their overlapping portions to assemble the sequences of larger DNA regions and later, entire chromosomes. This process is still popularly used today to sequence individual DNA pieces used for cloning or through PCR.

Next-generation sequencing Next-gen sequencing is like running multiple small sanger sequencing reactions at the same time. This allows sequencing to be faster and cheaper than Sanger.