IB Biology SL Topic D1.1

What is semi-conservative replication?

Semi-conservative replication is the method by which DNA copies itself. During replication, the original double-stranded DNA molecule unwinds and separates, and each of the two single strands acts as a template for a new strand. As a result, each of the two newly formed DNA molecules consists of one original (parental) strand and one newly synthesized strand.

Which scientists famously carried out an experiment to determine the method of replication?

Matthew Meselson and Franklin Stahl carried out the famous 1958 experiment using isotopes of nitrogen and density gradient centrifugation, which proved that DNA replication is semi-conservative.

What feature of DNA allows for accurate replication?

Complementary base pairing. Because adenine (A) always pairs with thymine (T), and cytosine (C) always pairs with guanine (G) via hydrogen bonds, each template strand dictates the exact sequence of the new strand.

Draw and annotate DNA replication, including these details: helicase, DNA polymerase, direction of replication, free nucleotides, complementary base pairing.

• Helicase: Unzips the double helix by breaking the hydrogen bonds between complementary base pairs at the replication fork.

• DNA polymerase: The enzyme that links free nucleotides together to form the new complementary strand.

• Direction of replication: Synthesis always occurs in the 5' to 3' direction on the new strand.

• Free nucleotides: Deoxynucleoside triphosphates floating in the nucleoplasm that are brought in to match template bases.

• Complementary base pairing: Ensures exact copies are made (A-T and C-G).

What does PCR stand for?

Polymerase Chain Reaction.

What specific DNA polymerase is used in PCR, and why?

Taq polymerase (extracted from the thermophilic bacterium Thermus aquaticus) is used. It is used because it is heat-stable and does not denature (break down) at the high temperatures ($95^\circ\text{C}$) required to separate DNA strands during PCR.

Describe the first step of PCR: denaturation.

The DNA sample is heated to roughly $94^\circ\text{C}$–$98^\circ\text{C}$. This high temperature breaks the hydrogen bonds holding the two DNA strands together, causing the double helix to separate into two single strands.

Describe the second step of PCR: annealing.

The mixture is cooled to around $50^\circ\text{C}$–$65^\circ\text{C}$. This allows short, single-stranded DNA sequences called primers to bind (anneal) to complementary sequences on the separated DNA templates, marking the starting point for replication.

Describe the third step of PCR: elongation.

The temperature is raised to around $72^\circ\text{C}$ (the optimal operating temperature for Taq polymerase). Taq polymerase attaches free nucleotides to the primer, extending the new DNA strand in the 5' to 3' direction.

Why is PCR exponential?

Each thermal cycle (denaturation → annealing → elongation) doubles the number of DNA molecules. One molecule becomes 2, 2 become 4, 4 become 8, and so on, resulting in exponential amplification (2^n, where n is the number of cycles).

How are the DNA samples amplified and made into fragments?

DNA is first amplified using PCR to yield enough material. Then, restriction endonucleases (restriction enzymes) are used to cut the DNA strands at specific, predictable recognition sites, creating DNA fragments of varying lengths.

Why are short tandem repeats of interest in DNA profiling?

Short Tandem Repeats (STRs) are short sequences of DNA that repeat over and over. They are highly polymorphic (vary greatly in length between individuals), making them ideal for creating unique DNA profiles to distinguish people.

Why do short tandem repeats exist, and why are they not removed from the genome?

STRs exist as repetitive elements within non-coding regions of DNA (sometimes referred to as satellite DNA). They are not removed because they do not code for proteins; thus, mutations or variations in their length generally have no negative impact on an organism's survival or gene expression, meaning there is no selective pressure to eliminate them.

What technology is used to separate the DNA fragments by size?

Gel electrophoresis.

Why does DNA move through the gel in gel electrophoresis?

DNA has a net negative charge due to the phosphate groups in its backbone. When an electrical current is applied across the gel, the negatively charged DNA fragments are repelled by the negative electrode (cathode) and migrate toward the positive electrode (anode).

Which fragments will move quicker, and which will move slower, during gel electrophoresis?

• Smaller fragments move quicker and travel further through the pores of the gel matrix.

• Larger fragments move slower and stay closer to the starting wells.

In a crime scene case, how many fragments should match the profile of the suspect?

All of the detectable STR fragments/bands of the suspect must precisely match the DNA profile found at the crime scene for a positive identification.

In a paternity case, how many fragments should match the profile of the real father?

Half (50%) of the child's STR fragments must match the mother, and the remaining half (50%) must match the biological father (since a child inherits half of their genetic material from each parent).