DNA

A DNA mononucleotide is made of three components joined together by condensation.

The first is a phosphate group, which is bonded to carbon 5 of the sugar. The second is a pentose (5-carbon) sugar, deoxyribose. The third is one organic (nitrogenous) base, bonded to carbon 1 of the sugar; in DNA this base is one of adenine, thymine, cytosine or guanine.

These three parts together form the single repeating unit — the monomer — from which the polynucleotide strands of DNA are built.

In DNA, adenine always pairs with thymine, joined by two hydrogen bonds, and cytosine always pairs with guanine, joined by three hydrogen bonds. This is known as complementary base pairing.

Adenine and guanine are purines (larger, double-ring bases), while thymine and cytosine are pyrimidines (smaller, single-ring bases). Each pair therefore contains one purine bonded to one pyrimidine.

Because a large purine always pairs with a small pyrimidine, every base pair (or "rung" of the ladder) is the same width. This keeps the two sugar–phosphate backbones a constant distance apart along the whole length of the molecule, giving the double helix a regular, stable shape.

Each polynucleotide strand has a sugar–phosphate backbone in which the deoxyribose of one nucleotide is joined to the phosphate of the next by phosphodiester bonds, with the organic bases projecting from the backbone.

Two such strands lie alongside each other and are held together by hydrogen bonds between complementary bases — adenine with thymine (two hydrogen bonds) and cytosine with guanine (three hydrogen bonds).

The two strands run in opposite directions; they are described as antiparallel (one runs 5′ → 3′, the other 3′ → 5′). The bases face inwards towards the centre of the molecule.

The two strands are then twisted around a common axis to form a double helix.

Although each hydrogen bond is weak, there are a very large number of them along the molecule, so collectively they hold the two strands firmly together while still allowing the strands to be separated during replication.

Semi-conservative replication means that each of the two new DNA molecules produced is built from one strand of the original (parent) molecule and one newly synthesised strand. The original strand is therefore conserved in each new molecule, which is why the original genetic information is passed on accurately.

The enzyme helicase moves along the molecule and breaks the hydrogen bonds between the complementary bases, unwinding and separating the two strands so that each can act as a template.

Free DNA mononucleotides then line up against each template strand by complementary base pairing. The enzyme DNA polymerase catalyses the condensation reactions that join these nucleotides together, forming the phosphodiester bonds of the new sugar–phosphate backbone.

The result is two identical DNA molecules, each consisting of one old strand and one new strand.

Growing the bacteria in $^{15}N$ for many generations meant that all the original DNA was heavy, because nitrogen is part of every base. After transfer to $^{14}N$, any newly made strands are built from light nitrogen, so density reveals how old and new strands are distributed.

After one round of replication a single band of intermediate density appears. Under the semi-conservative model each molecule contains one heavy original strand and one light new strand, so every molecule has the same intermediate density — exactly one band. This result rules out the conservative model, in which the original heavy double helix would be kept intact and a completely new light double helix made, producing one heavy band and one light band rather than a single intermediate band.

After two rounds of replication there are two bands, one intermediate and one light, in equal amounts. The heavy original strands made in the first generation persist, each now paired with a new light strand to give intermediate molecules; the remaining molecules are made of two new light strands and are fully light. This 1:1 intermediate-to-light pattern fits the semi-conservative model precisely.

The appearance of a distinct light band at the second generation rules out the dispersive model, in which old and new nucleotides are scattered through both strands of every molecule. The dispersive model predicts a single band that becomes progressively lighter each generation but never splits into two separate bands.

Because only the semi-conservative model — one conserved strand plus one new strand — accounts for both the single intermediate band at generation one and the intermediate-plus-light bands at generation two, the experiment provides strong evidence for it.

(a) Because cytosine always pairs with guanine, the percentage of cytosine equals the percentage of guanine, so cytosine = 32%. Guanine and cytosine together therefore account for $32 + 32 = 64\%$ of the bases.

The remaining percentage must be made up of adenine and thymine: $100 - 64 = 36\%$. Because adenine always pairs with thymine, this 36% is shared equally, so adenine = 18% and thymine = 18%.

So the composition is: guanine 32%, cytosine 32%, adenine 18%, thymine 18% (which correctly totals 100%).

(b) In double-stranded DNA, each base on one strand is paired with its complementary base on the other strand: adenine with thymine and cytosine with guanine. This means the amount of adenine always equals the amount of thymine, and the amount of cytosine always equals the amount of guanine (Chargaff's rule).

Therefore, knowing one base percentage immediately gives its partner (here C = G = 32%). Since all four percentages must add up to 100%, the remaining percentage is fixed and is shared equally between the other complementary pair (A = T = 18%). It is the strict one-to-one complementary base pairing that makes this deduction possible.

Adjacent nucleotides in a single DNA strand are joined by a phosphodiester bond. This covalent bond links the phosphate group on carbon 5′ of one nucleotide to the hydroxyl group on carbon 3′ of the next nucleotide.

The bond is formed by a condensation reaction, in which a molecule of water is removed each time a bond is made.

The presence of ribose as the pentose sugar and uracil as a base shows that this nucleic acid is RNA, not DNA.

Three further differences between DNA and RNA are:

• Pentose sugar: DNA contains deoxyribose (one fewer oxygen atom), whereas RNA contains ribose.
• Bases: DNA uses the base thymine, whereas RNA uses uracil in its place; both still use adenine, cytosine and guanine.
• Number of strands / length: DNA is a double-stranded molecule (two antiparallel polynucleotide strands in a double helix) and is typically very long, whereas RNA is usually a single, shorter polynucleotide strand.

In RNA, adenine pairs with uracil rather than thymine, but the principle of complementary base pairing (a purine with a pyrimidine, held by hydrogen bonds) is the same.

The two strands of a DNA molecule are held together by hydrogen bonds between complementary bases — adenine with thymine, cytosine with guanine. Because this base pairing is complementary, the base sequence of one strand exactly determines the sequence of the other.

During replication, the enzyme helicase breaks the hydrogen bonds between the bases, separating the two strands so that each acts as a template. The covalent phosphodiester bonds of the sugar–phosphate backbone are not broken, so the base sequence of each template strand is preserved intact.

Free DNA mononucleotides then line up against each template by complementary base pairing: a free nucleotide can only bond where its base is complementary to the template base. The enzyme DNA polymerase catalyses condensation reactions that join these nucleotides by phosphodiester bonds, building each new strand.

Because each new strand is assembled by complementary base pairing onto a conserved template, the sequence of the new molecule must match that of the original. Replication is semi-conservative — each new molecule contains one original strand and one new strand — so the original strand acts as a faithful guide. As a result, the two daughter cells each receive DNA carrying an identical base sequence, conserving the genetic information.

Trend in the data. As the generations increase, the heavy band disappears completely after generation 0 (100% → 0%). The intermediate band rises to 100% at generation 1 then halves each generation (100% → 50% → 25%), while the light band rises steadily (0% → 50% → 75%). This shows old (heavy) and new (light) strands are being redistributed in a regular pattern.

Generation 0 → 1: the heavy band vanishes and 100% is intermediate. Every molecule now contains one original ¹⁵N strand and one new ¹⁴N strand, giving uniform intermediate density. This rules out the conservative model: if the parent double helix were kept fully intact and an entirely new double helix made, generation 1 would show 50% heavy and 50% light bands and no intermediate band — which is not seen.

Generation 2: a distinct light band appears (50% intermediate, 50% light). The two original ¹⁵N strands persist, each paired with a new ¹⁴N strand to give the 50% intermediate; the remaining molecules are built from two new ¹⁴N strands and form the 50% light band. The appearance of two separate bands at this stage rules out the dispersive model: dispersive replication scatters old and new nucleotides through every strand, so it predicts a single band of steadily decreasing (uniform) density that never splits into two.

Generation 3 confirms the pattern. Under the semi-conservative model the two conserved ¹⁵N strands always remain as intermediate molecules, while the proportion of fully-light DNA grows. The predicted intermediate fraction is $\dfrac{2}{2^{3}} = \dfrac{2}{8} = 25\%$, exactly matching the data (25% intermediate, 75% light). The conserved heavy strands become a smaller and smaller fraction as the population grows exponentially.

Because only the semi-conservative model — one strand conserved, one strand new — predicts both the single intermediate band at generation 1 and the splitting into intermediate-plus-light bands that always retains a halving intermediate fraction, the data provide strong evidence for it and eliminate the conservative and dispersive alternatives.