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Educerie · IB Diploma · Biology
Theme D Continuity and change · D1.1 DNA replication
What you must be able to do
| You must be able to | Level | What it looks like in the exam |
|---|---|---|
| State why DNA must be replicated, and that replication makes exact copies | SL, HL | Paper 1A item |
| Explain semi-conservative replication and how complementary base pairing makes it accurate | SL, HL | "Explain how the semi-conservative nature of replication…" (3 marks) |
| Outline the roles of helicase and DNA polymerase | SL, HL | "Outline the roles of two enzymes in DNA replication" (2 marks) |
| Explain PCR: primers, the three temperatures, Taq polymerase | SL, HL | "Explain the role of Taq polymerase" (2 marks); a copies calculation |
| Explain how gel electrophoresis separates DNA fragments | SL, HL | "Explain why the fragments separate" (2 marks) |
| Interpret DNA profiles for paternity and forensics, and explain why more markers make a match more reliable | SL, HL | Paper 1B gel or profile, 3–4 marks |
| Explain the 5′ to 3′ directionality of DNA polymerases | HL only | "Explain why DNA replication is 5′ to 3′" (2 marks) |
| Distinguish replication on the leading and lagging strands, with Okazaki fragments | HL only | "Distinguish between…" (3–4 marks) |
| State the functions of primase, DNA polymerase III, DNA polymerase I and ligase | HL only | Paper 1A, or the core of an 8-mark Section B answer |
| Explain proofreading by DNA polymerase III | HL only | Short answer, 2 marks |
Before you start
You need A1.2 in full: the nucleotide, the sugar–phosphate backbone, the double helix of two antiparallel strands, and complementary base pairing, A with T and C with G, held by hydrogen bonds. HL students also need the 5′ and 3′ ends of a strand from A1.2's HL section. From C1.1 you need the idea of an enzyme with a specific job. The only maths is doubling.
1The idea in one paragraph
Before a cell divides, it copies all of its DNA so that each daughter cell gets a complete set. Helicase unzips the double helix by breaking the hydrogen bonds between the bases, and each exposed strand becomes a template. Free nucleotides pair with the template bases, A with T and C with G, and DNA polymerase links them into a new strand. Because each base has only one partner, the new strand is exactly complementary to its template, and each new molecule is one old strand plus one new one: semi-conservative replication. The same chemistry, run in a tube, is the polymerase chain reaction, which turns a trace of DNA into billions of copies, and gel electrophoresis then sorts the copies by length, which is how DNA profiles settle paternity and solve crimes. At HL the detail matters: polymerase can only build 5′ to 3′, so one new strand is built smoothly and the other in short pieces, with four more enzymes to start, finish, join and check them.
2Why DNA is replicated
DNA replication is the production of exact copies of DNA, with base sequences identical to the original. It happens before every cell division, and cell division is needed for three things.
- Reproduction. A single-celled organism reproduces by dividing, and a sexually reproducing organism makes gametes by division. Each new cell needs a full copy of the genetic information.
- Growth. A multicellular organism grows by adding cells. You grew from one cell to tens of trillions, and every one received a copy of your DNA.
- Tissue replacement. Cells wear out and are replaced all through life: skin cells, the lining of the gut, red blood cells made in bone marrow. Each replacement cell needs its DNA.
"Exact" is the key word. A copy with a changed base sequence could code for a faulty protein. The mechanism in section 3 is what makes exact copying possible.
3Semi-conservative replication and complementary base pairing
When DNA is replicated, the two strands of the double helix separate, and each old strand acts as a template for building a new one. Figure 1 follows eight base pairs through the process.
A template works because of complementary base pairing. Opposite every A on the template, only a T fits; opposite every C, only a G; and the other way round. So the base sequence of the template fixes the base sequence of the new strand completely. The top strand in Figure 1 reads ATGCCGTA, so the strand built on it must read TACGGCAT, which is exactly the sequence of the strand it was separated from.
Each of the two new DNA molecules therefore contains one strand from the parent molecule and one newly made strand. That is what semi-conservative means: half of each original molecule is conserved in each copy.
DNA replication is semi-conservative: each new molecule is one original strand, used as a template, paired with one new complementary strand.
Why does this give a high degree of accuracy? Two reasons work together.
- Each base has one partner. Pairing depends on the shapes of the bases and on hydrogen bonds that only form between A and T or between C and G. A wrong base does not fit well and is unlikely to be added.
- Each strand carries the whole message. Because the two strands are complementary, either strand alone contains all the information needed to rebuild the other. Nothing is copied from memory or guesswork; every new base is chosen by the template base opposite it.
Mistakes still happen, but very rarely. HL section 10 describes one of the ways cells catch the few that slip through.
4Helicase and DNA polymerase
Two enzymes do the main work, and Figure 2 shows where each acts.
Helicase unwinds the double helix and separates the two strands by breaking the hydrogen bonds between complementary bases. It moves along the DNA, opening it up like a zip, and the Y-shaped region where the strands are coming apart is called the replication fork. Helicase breaks only hydrogen bonds; the covalent bonds of the sugar–phosphate backbone stay intact, so each template strand remains whole.
DNA polymerase builds the new strands. Free DNA nucleotides in the nucleus (or in the cytoplasm of a prokaryote) pair with the exposed bases of the template by complementary base pairing, and DNA polymerase joins each new nucleotide to the end of the growing strand with a covalent bond, forming the new sugar–phosphate backbone. The guide limits you at SL to this general role: DNA polymerase assembles a new strand, one nucleotide at a time, complementary to the template.
A common slip is to say DNA polymerase "breaks" or "unzips" anything. It does not. Helicase separates; polymerase builds.
5The polymerase chain reaction: copying DNA in a tube
Biologists often have only a trace of DNA: a spot of blood, a hair root, a few cells from a swab. That is far too little to analyse. The polymerase chain reaction (PCR) copies a chosen section of DNA over and over, so that a few molecules become billions. It is DNA replication run in a machine, with temperature doing the jobs that helicase and the cell's controls do inside a cell. Figure 3 shows one cycle.
The tube contains the DNA sample, a large supply of free nucleotides, two kinds of primer, and Taq polymerase, in a buffer. A machine called a thermal cycler changes the temperature in a repeated cycle.
- Denaturation, about 95 °C. The heat breaks the hydrogen bonds between the strands, so the DNA separates into single strands. This does the job of helicase.
- Annealing, about 55 °C. The mixture is cooled so that primers can bind (anneal) by complementary base pairing to the single strands. A primer is a short, single-stranded length of DNA, about 20 nucleotides long, designed to match the sequence at one end of the target section. There are two primers, one for each strand, and together they mark the start and end of the section to be copied. The polymerase needs a primer to start from, so only the section between the primers is copied.
- Extension, about 72 °C. Taq polymerase adds nucleotides to each primer, building a new strand complementary to each template. From one double-stranded molecule, there are now two.
Why Taq polymerase? It is a DNA polymerase from Thermus aquaticus, a bacterium that lives in hot springs. Its enzymes are adapted to high temperatures, so it is not denatured at 95 °C and works best at around 72 °C. A human DNA polymerase would be denatured in the first heating step and would have to be replaced every cycle.
Each cycle doubles the number of copies, so the number grows exponentially:
Thirty cycles take a couple of hours and turn a single molecule into about a billion copies of the target sequence.
6Gel electrophoresis: sorting DNA by length
Once DNA has been copied, the fragments must be separated so they can be compared. Gel electrophoresis separates DNA fragments by length, as Figure 4 shows.
The DNA samples are placed in small wells at one end of a slab of gel, a jelly-like material full of microscopic pores. The gel is covered in a buffer solution and an electric current is passed through it. The basis of the separation is two facts.
- DNA is negatively charged, because of the phosphate groups in its backbone. So all DNA fragments move through the gel towards the positive electrode.
- Short fragments move faster than long ones, because they pass more easily through the pores of the gel. After a set time, the shortest fragments have travelled furthest.
The fragments end up as bands, each band being many copies of a fragment of one length. They are made visible with a stain. A DNA ladder, a mixture of fragments of known lengths, is run in one lane so that the length of any band can be read off by comparing its position. In Figure 4, sample 1 contains fragments of about 700 and 300 base pairs (bp).
7Applications of PCR and gel electrophoresis
PCR and gel electrophoresis are used together almost everywhere DNA is studied: to test for an infection by detecting a pathogen's genetic material, to check whether a crop contains a genetically modified gene, to identify the species of a fish sold in a market, to study DNA from ancient bones. The two the guide names are DNA profiling for paternity and for forensic investigations.
How a DNA profile works. Everyone's DNA contains regions where a short sequence of bases is repeated many times in a row. The number of repeats at each such region varies from person to person, so the length of the region varies too. Each of these variable regions is a marker. A person inherits one version of each marker from each parent, so they have two lengths at each marker (or one, if both happen to be the same). PCR copies several markers from a sample, and electrophoresis separates the copies by length. The resulting pattern of bands is a DNA profile.
Paternity. Every band in a child's profile must have come from either the mother or the biological father. Figure 5 shows the logic.
Match the child's bands to the mother's first. The bands left over (amber) must have come from the father. Man B has both of them; man A has neither, so man A is excluded, and the profile is consistent with man B being the father.
Forensics. DNA from a crime scene, from blood, skin cells or hair roots, is amplified by PCR and profiled, then compared with the profiles of suspects or with a database. A suspect whose profile differs at any marker is excluded. A suspect whose profile matches at every marker is very probably the source, though a match shows only that the DNA is theirs, not how it got there.
Nature of science: reliability and the number of markers. In any experiment, reliability improves as the number of measurements increases. In DNA profiling, each marker is a separate measurement. Suppose, for illustration only, that any one marker's pair of lengths is shared by 1 person in 10. Then:
Each extra marker multiplies down the probability of a false match, where an innocent person's profile matches by chance. That is why forensic laboratories do not use one or two markers: national DNA databases use sets of around sixteen to twenty.
8HLDirectionality of DNA polymerases
SL students can skip to section 11.
Each strand of DNA has two different ends. From A1.2: the carbons of deoxyribose are numbered 1′ to 5′, the phosphate is attached to carbon 5′, and when nucleotides are linked the phosphate of one bonds to carbon 3′ of the next. So one end of a strand, the 5′ end, has a free phosphate on a 5′ carbon, and the other end, the 3′ end, has a free –OH group on a 3′ carbon.
DNA polymerases can only add nucleotides to the 3′ end of a strand. The incoming nucleotide's 5′ phosphate is bonded to the free 3′ –OH at the end of the growing strand. So every new strand is built in the 5′ to 3′ direction. Figure 6 shows the rule.
Because the new strand is antiparallel to its template, polymerase moves along the template from the template's 3′ end towards its 5′ end. Remember it in one line: DNA polymerase adds the 5′ of a new nucleotide to the 3′ end of the strand.
9HLThe leading strand, the lagging strand and the enzymes that build them
The 5′ to 3′ rule creates a problem at the replication fork. The two template strands are antiparallel, and helicase opens both of them in the same direction. So one new strand can be built in the same direction as the fork is moving, and the other must be built in the opposite direction. Figure 7 shows how the cell solves it, for a prokaryote such as E. coli.
The leading strand is built continuously, towards the fork. As helicase opens more template, polymerase simply keeps adding nucleotides to the same 3′ end. It needs to be started only once.
The lagging strand is built discontinuously, away from the fork, in short pieces called Okazaki fragments. Each time helicase exposes a new stretch of template, a new fragment is started close to the fork and built backwards, 5′ to 3′, until it reaches the fragment made before it. The fragments are then joined. Because every fragment needs its own start, the lagging strand must be started repeatedly.
DNA polymerase cannot start a strand from nothing; it can only add to an existing 3′ end. So every new strand begins with a short RNA primer, and four enzymes besides helicase do the work. The guide limits this to the prokaryotic system.
| Enzyme | Function |
|---|---|
| DNA primase | Makes a short RNA primer, complementary to the template, which gives DNA polymerase III a 3′ end to build from. Once on the leading strand; at the start of every Okazaki fragment on the lagging strand. |
| DNA polymerase III | Adds DNA nucleotides to the 3′ end of the primer, building the new strand 5′ to 3′: continuously on the leading strand, one fragment at a time on the lagging strand. Also proofreads (section 10). |
| DNA polymerase I | Removes the RNA primer nucleotides and replaces them with DNA nucleotides. |
| DNA ligase | Seals the remaining gap in the sugar–phosphate backbone between adjacent fragments by making the final covalent bond, joining the Okazaki fragments into one continuous strand. |
So the order of events on the lagging strand is: primase lays a primer, DNA polymerase III extends it into an Okazaki fragment, DNA polymerase I swaps the primer for DNA, and DNA ligase joins the fragment to its neighbour.
| Leading strand | Lagging strand | |
|---|---|---|
| Direction of synthesis relative to the fork | towards the fork | away from the fork |
| How it is made | continuously | discontinuously, as Okazaki fragments |
| RNA primers needed | one, once | one for every fragment, repeatedly |
| DNA ligase needed | not for joining fragments | yes, to join every fragment |
| Direction of synthesis | 5′ to 3′ | also 5′ to 3′ |
The last row is the one students forget: both strands are built 5′ to 3′. The difference is only in whether that direction points towards the fork or away from it.
10HLProofreading
DNA polymerase III does occasionally add a nucleotide whose base does not pair with the template. It then corrects its own mistake, and this is proofreading, shown in Figure 8.
Before adding the next nucleotide, DNA polymerase III checks the base at the 3′ terminal of the strand it is building. If that base is mismatched, such as a G opposite a T, it does not fit properly, and polymerase III removes that nucleotide from the 3′ end. It then replaces it with the correctly matched nucleotide, an A opposite the T, and carries on. Proofreading catches most of the errors that complementary base pairing lets through, which is a large part of why replication is so accurate.
11Where marks are lost
Saying helicase breaks the backbone, or "breaks the DNA". Helicase breaks the hydrogen bonds between bases; the covalent backbone is untouched.
Swapping the jobs of the two enzymes. Helicase unwinds and separates; DNA polymerase links nucleotides into the new strand. Polymerase does not unzip anything.
Describing replication as "conservative" or "the old DNA is copied into a new molecule". Semi-conservative means each new molecule has one old strand and one new strand.
Thinking DNA moves towards the negative electrode. DNA is negatively charged (phosphates), so it moves towards the positive electrode, and the shortest fragments move furthest.
Saying Taq polymerase is used because it "works faster" or "is not affected by heat". The point is that it comes from a hot-spring bacterium and is not denatured at the 95 °C of the denaturation step, so it survives every cycle.
Saying a DNA match "proves guilt" or that one marker is enough. A match shows the DNA is probably the suspect's; more markers reduce the chance of a false match.
HL: saying the lagging strand is built 3′ to 5′. Both strands are built 5′ to 3′. The lagging strand is built away from the fork, in fragments.
HL: giving DNA polymerase I's job to ligase, or the reverse. Polymerase I replaces RNA primers with DNA; ligase seals the final gap between fragments.
12Draw it right
You may be asked to draw semi-conservative replication, a replication fork, a gel, or to annotate one. Markers look for the following.
- Two strands in each molecule, with old and new strands distinguished (shading, colour or a key).
- Semi-conservative result: two daughter molecules, each with one old and one new strand, and the base sequences complementary (A–T, C–G) and identical between the two daughters.
- Helicase at the fork, labelled with its function: breaking hydrogen bonds between bases.
- Gel: wells at the negative end, bands further towards the positive end for shorter fragments, a ladder lane labelled with sizes, and the direction of movement shown.
- HL fork: templates labelled 5′ and 3′ at their ends; leading strand continuous towards the fork; lagging strand as several Okazaki fragments built away from it; RNA primers at the 5′ end of the leading strand and of each fragment; primase, polymerase III, polymerase I and ligase each labelled where they act.
- HL arrows: any arrow showing synthesis points in the 5′ to 3′ direction of the new strand.
13Try it
Marks in brackets. Answers and marker's notes are at the end.
Q1. Explain how complementary base pairing and the semi-conservative nature of replication allow DNA to be copied accurately. 4 marks
Q2. Outline the roles of helicase and DNA polymerase in DNA replication. 2 marks
Q3. A forensic scientist uses PCR to amplify DNA from a small bloodstain.
(a) State the role of the primers. 1 mark
(b) Explain why Taq polymerase is used rather than a human DNA polymerase. 2 marks
(c) The sample contains 5 copies of the target sequence. Calculate the number of copies after 12 cycles, assuming every cycle doubles the number. 1 mark
Q4. DNA from a crime scene and from three suspects was profiled at three markers. The table gives the fragment lengths, in base pairs, found at each marker. The data are invented.
| Marker | Crime scene | Suspect 1 | Suspect 2 | Suspect 3 |
|---|---|---|---|---|
| M1 | 180, 204 | 180, 204 | 180, 204 | 176, 212 |
| M2 | 232, 244 | 228, 244 | 232, 244 | 232, 240 |
| M3 | 150, 162 | 150, 158 | 150, 162 | 146, 162 |
(a) Identify the suspect whose DNA matches the crime scene sample. 1 mark
(b) Using suspect 1, explain why profiles are compared at several markers rather than one. 2 marks
(c) On a gel, state which of the two fragments at marker M1 would travel further, and why. 2 marks
Q5 (HL). Explain the differences between DNA replication on the leading strand and on the lagging strand. 5 marks
Q6 (HL). Outline how DNA polymerase III proofreads a newly synthesised strand. 2 marks
14In one breath
DNA is replicated before every division, for reproduction, growth and tissue replacement, and the copies must be exact. Helicase unwinds the helix and breaks the hydrogen bonds between bases; each old strand is a template; free nucleotides pair A–T and C–G; DNA polymerase links them into a new strand. Each new molecule is one old strand and one new one: semi-conservative, and accurate because each base has only one partner. PCR copies a chosen section in a tube: 95 °C to separate strands, about 55 °C for primers to anneal to each end of the target, 72 °C for heat-stable Taq polymerase to extend them, doubling every cycle, 2ⁿ. Gel electrophoresis sorts fragments by length: DNA is negative, runs to the positive electrode, shortest furthest, sized against a ladder. DNA profiles compare variable-length markers: every band in a child comes from the mother or the father, and more markers mean less chance of a false match. HL: polymerase adds only to the 3′ end, so strands grow 5′ to 3′; the leading strand is continuous with one primer, the lagging strand discontinuous in Okazaki fragments, each with its own primer; primase lays RNA primers, polymerase III extends them and proofreads by removing a mismatched 3′ nucleotide, polymerase I swaps primers for DNA, and ligase seals the gaps.
Answers
Q1. The two strands of the DNA double helix separate and each acts as a template for a new strand. Free nucleotides pair with the exposed bases by complementary base pairing, A with T and C with G, since only these pairs form hydrogen bonds with each other. So the sequence of each new strand is determined exactly by its template, and is identical to the strand that was originally paired with that template. Each new molecule consists of one original strand and one new strand (semi-conservative), so both daughter molecules have base sequences identical to the parent molecule. 1 for strands separating and each acting as a template, 1 for the pairing rules, 1 for the template determining the new sequence exactly, 1 for one old plus one new strand giving two identical molecules. "Semi-conservative" stated without explanation scores 0 for that point.
Q2. Helicase unwinds the double helix and separates the strands by breaking the hydrogen bonds between complementary bases. DNA polymerase links free nucleotides together (with covalent bonds) to form a new strand complementary to the template strand. 1 for each enzyme. Helicase "unzips the DNA" with no mention of hydrogen bonds is accepted only if separation of strands is clear.
Q3. (a) Primers bind to the ends of the target sequence and provide a starting point from which the polymerase can add nucleotides. (b) Taq polymerase comes from a bacterium adapted to hot springs, so it is not denatured at the high temperature (about 95 °C) used to separate the strands in each cycle; a human DNA polymerase would be denatured and would need replacing after every cycle. (c) 5 × 2¹² = 5 × 4,096 = 20,480 copies. (a) 1. (b) 1 for not denatured at high temperature, 1 for linking this to the denaturation step of every cycle. (c) 1 for 20,480. An answer of 2¹² = 4,096 ignores the starting copies and scores 0 in (c).
Q4. (a) Suspect 2. (b) Suspect 1 matches the crime scene sample at marker M1 (180, 204) but not at M2 (228 instead of 232) or M3 (158 instead of 162). If only M1 had been used, suspect 1 would have been wrongly matched. Each additional marker makes a chance match by an innocent person less likely, so using several markers makes the conclusion more reliable. (c) The 180 bp fragment, because shorter fragments move more easily through the pores of the gel, so they travel faster and further towards the positive electrode. (a) 1. (b) 1 for showing suspect 1 matches at one marker only, 1 for linking more markers to a lower probability of a false match. (c) 1 for 180 bp, 1 for the reason. Naming suspect 1 in (a) scores 0 and signals the student compared one marker.
Q5 (HL). DNA polymerase can only add nucleotides to the 3′ end of a strand, so both new strands are built 5′ to 3′; because the templates are antiparallel, one new strand is built towards the replication fork and one away from it. The leading strand is built continuously towards the fork, whereas the lagging strand is built discontinuously, away from the fork, as Okazaki fragments. The leading strand needs an RNA primer only once, at the start, whereas the lagging strand needs a new primer, made by primase, for every fragment. On the lagging strand DNA polymerase I then replaces each RNA primer with DNA, and DNA ligase joins the fragments by sealing the gaps in the sugar–phosphate backbone, whereas the leading strand does not need its fragments joined. 1 for 5′ to 3′ synthesis and antiparallel templates as the cause, 1 for continuous versus discontinuous, 1 for Okazaki fragments, 1 for one primer versus repeated primers, 1 for polymerase I and/or ligase acting on the lagging strand. "The lagging strand is made 3′ to 5′" loses the first mark.
Q6 (HL). DNA polymerase III checks the base of the most recently added nucleotide at the 3′ end of the new strand. If it is mismatched with the template base, polymerase III removes that nucleotide and replaces it with one carrying the correct complementary base, before continuing. 1 for detecting and removing a mismatched nucleotide from the 3′ terminal, 1 for replacing it with the correctly paired nucleotide. Attributing proofreading to ligase or helicase scores 0.
Educerie · written from the published IB Diploma Programme Biology guide, first assessment 2025, section D1.1 DNA replication. Original text, examples and questions. Diagrams drawn by Educerie. Last reviewed 25 September 2026.
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