Helicase, Polymerase, PCR, and Gel Electrophoresis

What is the role of helicase in DNA replication?

Helicase unwinds and separates the double‑stranded DNA, creating a replication fork.

What is the role of single‑stranded binding (SSB) proteins?

SSB proteins prevent the separated DNA strands from re‑annealing.

What is the role of DNA polymerase in DNA replication?

DNA polymerase adds complementary nucleotides to the template strand and extends the new strand in the 5'‑3' direction.

Why must DNA strands be separated before replication?

To allow replication machinery to access each strand as a template.

What is PCR (polymerase chain reaction)?

A technique used to amplify large quantities of a specific DNA sequence from a very small sample.

How many copies can standard PCR produce after 30 cycles?

Over 1 billion copies.

How do primers function in PCR?

They bind to a specific DNA segment so only the targeted region gets copied.

Why was PCR useful for COVID‑19 testing?

Primers targeting COVID‑19 genes allowed those sequences to be amplified and detected.

What happens during the heat stage of PCR?

The DNA sample is heated to separate strands by breaking hydrogen bonds.

What happens during the annealing stage of PCR?

The sample is cooled so primers can attach to the target DNA sequence.

What happens during the extension stage of PCR?

The sample is heated to about 75°C for Taq polymerase to build a complementary strand.

Why was high temperature a challenge in early PCR?

Typical polymerases denature at high temperatures required for strand separation.

What organism provided a solution to this problem?

Thermus aquaticus, a bacterium that thrives in hot springs.

What is Taq polymerase?

A heat‑stable DNA polymerase isolated from Thermus aquaticus.

Why is Taq polymerase essential for PCR?

It withstands high temperatures and can repeatedly copy DNA during PCR cycles.

What can PCR‑amplified DNA be used for?

It can be separated and analysed using gel electrophoresis.

What is gel electrophoresis?

A technique that separates DNA fragments based on size, mass, and electric charge.

How are DNA samples prepared for gel electrophoresis?

They are cut with restriction enzymes and labeled with dye.

What causes DNA fragments to move through the gel?

An electric current pulls them through the gel matrix.

Why do smaller DNA fragments move faster?

They are less obstructed by the gel and travel more quickly.

What is the final result of gel electrophoresis?

A unique DNA fingerprint specific to the individual.