PCR & Gel Electrophoresis – BIOL 201 Lecture Notes

Vocabulary flashcards covering core terms from the lecture on PCR, DNA sequencing, and agarose gel electrophoresis.

DNA Sequencing

Laboratory process that determines the exact order of nucleotides in a DNA segment or entire genome.

Whole-Genome Sequencing

Sequencing approach that aims to read the complete DNA of an organism, not just selected regions.

Polymerase Chain Reaction (PCR)

In-vitro technique that mimics semiconservative DNA replication to exponentially amplify a specific DNA segment.

Semiconservative DNA Replication

Natural cellular mechanism in which each daughter DNA molecule contains one original and one newly synthesized strand.

Taq Polymerase

Heat-stable DNA polymerase from Thermus aquaticus used in PCR to synthesize DNA at high temperatures.

Synthetic Primer

Short, lab-made single-stranded DNA sequence that anneals to target DNA to initiate PCR amplification.

Denaturation (PCR)

First PCR step (≈95 °C) where heat breaks hydrogen bonds, separating double-stranded DNA.

Annealing (PCR)

Second PCR step (≈50–65 °C) in which primers bind to complementary sequences on single-stranded DNA.

Extension / Synthesis (PCR)

Third PCR step (≈72 °C) where Taq polymerase adds nucleotides to primers, copying the target DNA.

Exponential Amplification

PCR outcome in which DNA quantity doubles each cycle, producing 2ⁿ copies after n cycles.

DNA Helicase

Enzyme that unwinds DNA in cellular replication; replaced by high heat in PCR.

Agarose Gel Electrophoresis

Technique that separates DNA fragments by size as they migrate through a porous agarose gel in an electric field.

Agarose

Linear polysaccharide forming a gel matrix with variable pore sizes, acting as a molecular sieve for DNA.

Negative Charge of DNA

Property arising from phosphate backbone, causing DNA to migrate toward the positive electrode in electrophoresis.

Molecular Sieve

Function of agarose gel in which smaller DNA fragments move more easily through the pores than larger ones.

Ethidium Bromide

Fluorescent intercalating dye that binds DNA and emits orange light under UV, allowing visualization after electrophoresis.

Midori Green

Safer fluorescent dye that binds DNA and glows green under UV or blue light.

Pathogen-Specific Primers

Primer pairs designed to anneal only to DNA from a particular pathogen, enabling diagnostic PCR.

Control PCR

Parallel reaction using known pathogen DNA to confirm that primers and reagents function properly.

Insertion Mutation

Genetic alteration where extra nucleotides are added, producing a larger PCR fragment than normal.

Deletion Mutation

Genetic alteration where nucleotides are lost, producing a smaller PCR fragment than normal.

Huntington’s Disease

Autosomal‐dominant disorder caused by >35 CAG repeats in the HTT gene; detected by sizing PCR fragments.

CAG Repeat

Tri-nucleotide sequence whose copy number in HTT correlates with Huntington’s disease severity.

Paternity Testing (PCR)

Use of PCR-amplified genetic markers to compare a child’s paternal alleles with those of an alleged father.

Allele

Different version of a gene; PCR can distinguish alleles by fragment size or sequence.

Thermus aquaticus

Thermophilic bacterium living in hot springs; source of Taq polymerase.

Primase

Enzyme that makes RNA primers in cellular replication; replaced by synthetic DNA primers in PCR.

Semicolumn DNA Ladder

Mixture of DNA fragments of known sizes run alongside samples to estimate fragment lengths on a gel.

Clinical Specimen

Sample (blood, swab, tissue, etc.) collected from a patient for diagnostic testing like PCR.

Agarose Concentration

Percentage of agarose in gel; higher percentages resolve smaller fragments, lower percentages resolve larger ones.