chemistry

Maxam–Gilbert sequencing

Maxam–Gilbert sequencing is a chemistry topic covered in the lgStudy science library. This page brings together a partial reference excerpt, illustrations, worked examples, real-world applications and a short study plan, so you can understand Maxam–Gilbert sequencing rather than just read about it. In short: Maxam–Gilbert sequencing is a method of DNA sequencing developed by Allan Maxam and Walter Gilbert in 1976–1977. This method is based on nucleobase-specific partial chemical modification of DNA and subsequent cleavage of the DNA backbone at sites adjacent to the modified nucleotides.

Key takeaways

Maxam–Gilbert sequencing belongs to chemistry; place it in that map before memorising details.
Learn the definition first, then one example that makes the definition concrete.
Connect Maxam–Gilbert sequencing to a quantity you can measure, compute or draw — that is where exam questions come from.
Reproduce the core statement of Maxam–Gilbert sequencing from memory before moving on to harder problems.

Reference excerpt

Maxam–Gilbert sequencing is a method of DNA sequencing developed by Allan Maxam and Walter Gilbert in 1976–1977. This method is based on nucleobase-specific partial chemical modification of DNA and subsequent cleavage of the DNA backbone at sites adjacent to the modified nucleotides.

Maxam–Gilbert sequencing was the first widely adopted method for DNA sequencing, and, along with the Sanger dideoxy method, represents the first generation of DNA sequencing methods. Maxam–Gilbert sequencing is no longer in widespread use, having been supplanted by next-generation sequencing methods.

History Although Maxam and Gilbert published their chemical sequencing method two years after Frederick Sanger and Alan Coulson published their work on plus-minus sequencing, Maxam–Gilbert sequencing rapidly became more popular, since purified DNA could be used directly, while the initial Sanger method required that each read start be cloned for production of single-stranded DNA. However, with the improvement of the chain-termination method (see below), Maxam–Gilbert sequencing has fallen out of favour due to its technical complexity prohibiting its use in standard molecular biology kits, extensive use of hazardous chemicals, and difficulties with scale-up. Allan Maxam and Walter Gilbert’s 1977 paper “A new method for sequencing DNA” was honored by a Citation for Chemical Breakthrough Award from the Division of History of Chemistry of the American Chemical Society for 2017. It was presented to the Department of Molecular & Cellular Biology, Harvard University.

Procedure Maxam–Gilbert sequencing requires radioactive labeling at one 5′ end of the DNA fragment to be sequenced (typically by a kinase reaction using gamma-32P ATP) and purification of the DNA. Chemical treatment generates breaks at a small proportion of one or two of the four nucleotide bases in each of four reactions (G, A+G, C, C+T). For example, the purines (A+G) are depurinated using formic acid, the guanines (and to some extent the adenines) are methylated by dimethyl sulfate, and the pyrimidines (C+T) are hydrolysed using hydrazine. The addition of salt (sodium chloride) to the hydrazine reaction inhibits the reaction of thymine for the C-only reaction. The modified DNAs may then be cleaved by hot piperidine; (CH2)5NH at the position of the modified base. The concentration of the modifying chemicals is controlled to introduce on average one modification per DNA molecule. Thus a series of labeled fragments is generated, from the radiolabeled end to the first "cut" site in each molecule. The fragments in the four reactions are electrophoresed side by side in denaturing acrylamide gels for size separation. To visualize the fragments, the gel is exposed to X-ray film for autoradiography, yielding a series of dark bands each showing the location of identical radiolabeled DNA molecules. From presence and absence of certain fragments the sequence may be inferred.

Related methods This method led to the Methylation Interference Assay, used to map DNA-binding sites for DNA-binding proteins. An automated Maxam–Gilbert sequencing protocol was developed in 1994.

Illustrations

Worked examples

Example 1 — a first encounter with Maxam–Gilbert sequencing

Start with the simplest possible case. Write down what Maxam–Gilbert sequencing claims or describes in one sentence, then invent the smallest concrete situation in which that sentence is true. In chemistry, the smallest case is usually a single object, a single equation or a single measurement. Check that every symbol or term in your sentence has a meaning in that case.

Example 2 — changing one variable

Take the situation from Example 1 and change exactly one quantity: double it, halve it, or set it to zero. Predict what should happen to Maxam–Gilbert sequencing before you calculate. Comparing your prediction with the result is the fastest way to find out whether you understand the idea or only the words.

Example 3 — an exam-style question

Typical questions about Maxam–Gilbert sequencing ask you to (a) state it precisely, (b) apply it to given data, and (c) explain a limitation. Practise writing all three answers in under five minutes; the third part is what separates a full-mark answer from an average one.

Applications of Maxam–Gilbert sequencing

In research: Maxam–Gilbert sequencing appears in chemistry research whenever the underlying quantities have to be modelled precisely. Papers usually cite it as a starting assumption and then explore where it breaks down.
In technology and industry: Engineering practice reuses Maxam–Gilbert sequencing in design rules, simulations and safety margins. Knowing the idea lets you read a specification sheet and understand why the numbers look the way they do.
In the classroom: Maxam–Gilbert sequencing is common in secondary-school and first-year university syllabi. It links to neighbouring topics 1977 in biotechnology, DNA sequencing, DNA sequencing methods, so understanding it makes those chapters shorter.
In everyday life: Look for Maxam–Gilbert sequencing outside the textbook — in sport, cooking, traffic, electronics or the sky above you. An example you found yourself is remembered far longer than one you were given.

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How to study Maxam–Gilbert sequencing in 20 minutes

Read the reference excerpt below once, without taking notes.
Close the page and write down what Maxam–Gilbert sequencing means in your own words.
Compare your version with the excerpt and mark what you missed.
Work through the three examples above with pen and paper.
Explain Maxam–Gilbert sequencing out loud to somebody else — or to Teacher Smith in the lgStudy chat.

Frequently asked questions

What is Maxam–Gilbert sequencing in simple terms?

Why does Maxam–Gilbert sequencing matter?

Because it connects several chemistry ideas at once: it gives you a definition you can apply, a quantity you can calculate, and a way to check whether a result is plausible.

How should I study Maxam–Gilbert sequencing?

Read the excerpt, restate it from memory, then work through the examples and applications listed on this page. The five-step study plan above takes about twenty minutes.

What does this page cover?

It gives you a compact reference excerpt plus original lgStudy explanations, examples, applications and study material on Maxam–Gilbert sequencing.