chemistry

Molecular Inversion Probe

Molecular Inversion Probe 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 Molecular Inversion Probe rather than just read about it. In short: Molecular Inversion Probe (MIP) belongs to the class of Capture by Circularization molecular techniques for performing genomic partitioning, a process through which one captures and enriches specific regions of the genome. Probes used in this technique are single stranded DNA molecules and, similar to other genomic partitioning techniques, contain sequences that are complementary to the target in the genome; these p…

Key takeaways

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Connect Molecular Inversion Probe to a quantity you can measure, compute or draw — that is where exam questions come from.
Reproduce the core statement of Molecular Inversion Probe from memory before moving on to harder problems.

Reference excerpt

Molecular Inversion Probe (MIP) belongs to the class of Capture by Circularization molecular techniques for performing genomic partitioning, a process through which one captures and enriches specific regions of the genome. Probes used in this technique are single stranded DNA molecules and, similar to other genomic partitioning techniques, contain sequences that are complementary to the target in the genome; these probes hybridize to and capture the genomic target. MIP stands unique from other genomic partitioning strategies in that MIP probes share the common design of two genomic target complementary segments separated by a linker region. With this design, when the probe hybridizes to the target, it undergoes an inversion in configuration (as suggested by the name of the technique) and circularizes. Specifically, the two target complementary regions at the 5' and 3' ends of the probe become adjacent to one another while the internal linker region forms a free hanging loop. The technology has been used extensively in the HapMap project for large-scale SNP genotyping as well as for studying gene copy alterations and characteristics of specific genomic loci to identify biomarkers for different diseases such as cancer. Key strengths of the MIP technology include its high specificity to the target and its scalability for high-throughput, multiplexed analyses where tens of thousands of genomic loci are assayed simultaneously.

Technique Procedure

Molecular Inversion Probe Structure The probes are designed with sequences that are complementary to the genomic target at its 5' and 3' ends . The internal region contains two universal PCR primer sites that are common to all MIPs as well as a probe-release site, which is usually a restriction site. If the identification of the captured genomic target is performed using array-based hybridization approaches, the internal region may optionally contain a probe-specific tag sequence that uniquely identifies the given probe as well as a tag-release site, which, similar to the probe-release site, is also a restriction site.

Protocol Anneal probe to genomic target DNA Probes are added to the genomic DNA sample. After a denaturation followed by an annealing step, the target-complementary ends of the probe are hybridized to the target DNA. The probes then undergo circularization in this process. These probes, however, are designed such that a gap delimited by the hybridized ends of the probes remains over the target region. The size of the gap ranges from a single nucleotide for SNP genotyping to several hundred nucleotides for loci capture (e.g. exome capture).

Gap filling The gap is filled by DNA polymerase using free nucleotides and the ends of the probe are ligated by ligase, resulting in a fully circularized probe.

Remove non-reacted probes Since gap filling is not performed for non-reacted probes, they remain linear. Exonuclease treatment removes these non-reacted probes as well as any remaining linear DNA in the reaction.

Probe release In some versions of the protocol, the probe-release site (commonly a restriction site) is cleaved by restriction enzymes such that the probe becomes linearized. In this linearized probe the universal PCR primer sequences are located at the 5' and 3' ends and the captured genomic target becomes part of the internal segment of the probe. Other protocols leave the probe as a circularized molecule.

Captured target enrichment If the probe is linearized, traditional PCR amplification is performed to enrich the captured target using the universal primers of the probe. Otherwise, rolling circle amplification is performed for the circular probe.

Captured target identification The captured target can be identified either via array-based hybridization approaches or by sequencing of the target. If array-based approach is used, the probe may optionally contain a probe-specific tag that uniquely identifies the probe as well as the genomic region targeted by it. The tags from each probe are released by cleaving the tag release site with restriction enzymes. These tags are then hybridized to the sequences that are placed on the array and are complementary to them. The captured target can also be identified by sequencing the probe, now also containing the target. Traditional Sanger sequencing or cheaper, more high-throughput technologies such as SOLiD, Illumina or Roche 454 can be used for this purpose. Multiplex analysis Although each probe examines one specific genomic locus, multiple probes can be combined into a single tube for multiplexed assay that simultaneously examines multiple loci. Currently, multiplexed MIP analysis can examine more than 55,000 loci in a single assay.

Technique Development History

… excerpt ends here. Continue reading the full article.

Illustrations

Worked examples

Example 1 — a first encounter with Molecular Inversion Probe

Start with the simplest possible case. Write down what Molecular Inversion Probe 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 Molecular Inversion Probe 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 Molecular Inversion Probe 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 Molecular Inversion Probe

In research: Molecular Inversion Probe 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 Molecular Inversion Probe 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: Molecular Inversion Probe is common in secondary-school and first-year university syllabi. It links to neighbouring topics Genomics techniques, so understanding it makes those chapters shorter.
In everyday life: Look for Molecular Inversion Probe 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 Molecular Inversion Probe in 20 minutes

Read the reference excerpt below once, without taking notes.
Close the page and write down what Molecular Inversion Probe 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 Molecular Inversion Probe out loud to somebody else — or to Teacher Smith in the lgStudy chat.

Frequently asked questions

What is Molecular Inversion Probe in simple terms?

Why does Molecular Inversion Probe 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 Molecular Inversion Probe?

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 Molecular Inversion Probe.