chemistry

In silico PCR

In silico PCR 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 In silico PCR rather than just read about it. In short: In silico PCR refers to computational tools used to calculate theoretical polymerase chain reaction (PCR) results using a given set of primers (probes) to amplify DNA sequences from a sequenced genome or transcriptome. These tools are used to optimize the design of primers for target DNA or cDNA sequences.

Key takeaways

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

Reference excerpt

In silico PCR refers to computational tools used to calculate theoretical polymerase chain reaction (PCR) results using a given set of primers (probes) to amplify DNA sequences from a sequenced genome or transcriptome. These tools are used to optimize the design of primers for target DNA or cDNA sequences. Primer optimization has two goals: efficiency and selectivity. Efficiency involves taking into account such factors as GC-content, efficiency of binding, complementarity, secondary structure, and annealing and melting point (Tm). Primer selectivity requires that the primer pairs not fortuitously bind to random sites other than the target of interest, nor should the primer pairs bind to conserved regions of a gene family. If the selectivity is poor, a set of primers will amplify multiple products besides the target of interest.

The design of appropriate short or long primer pairs is only one goal of PCR product prediction. Other information provided by in silico PCR tools may include determining primer location, orientation, length of each amplicon, simulation of electrophoretic mobility, identification of open reading frames, and links to other web resources. Many software packages are available offering differing balances of feature set, ease of use, efficiency, and cost. Primer-BLAST is widely used, and freely accessible from the National Center for Biotechnology Information (NCBI) website. On the other hand, FastPCR, a commercial application, allows simultaneous testing of a single primer or a set of primers designed for multiplex target sequences. It performs a fast, gapless alignment to test the complementarity of the primers to the target sequences. Probable PCR products can be found for linear and circular templates using standard or inverse PCR as well as for multiplex PCR. Dicey is free software that outputs in-silico PCR products from primer sets provided in a FASTA file. It is fast (through use of a genome's FM-index) and can account for primer melting temperature and tolerated edit distances between primers and hit locations on the genome. VPCR runs a dynamic simulation of multiplex PCR, allowing for an estimate of quantitative competition effects between multiple amplicons in one reaction. The UCSC Genome Browser offers isPCR, which provides graphical as well text-file output to view PCR products on more than 100 sequenced genomes. A primer may bind to many predicted sequences, but only sequences with no or few mismatches (1 or 2, depending on location and nucleotide) at the 3' end of the primer can be used for polymerase extension. The last 10-12 bases at the 3' end of a primer are sensitive to initiation of polymerase extension and general primer stability on the template binding site. The effect of a single mismatch at these last 10 bases at the 3' end of the primer depends on its position and local structure, reducing the primer binding, selectivity, and PCR efficiency.

References

External links Webtools for PCR, qPCR, in silico PCR and oligonucleotides In silico simulation of molecular biology experiments

Illustrations

Worked examples

Example 1 — a first encounter with In silico PCR

Start with the simplest possible case. Write down what In silico PCR 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 In silico PCR 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 In silico PCR 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 In silico PCR

In research: In silico PCR 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 In silico PCR 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: In silico PCR is common in secondary-school and first-year university syllabi. It links to neighbouring topics Bioinformatics, Nucleic acids, so understanding it makes those chapters shorter.
In everyday life: Look for In silico PCR 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 In silico PCR in 20 minutes

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

Frequently asked questions

What is In silico PCR in simple terms?

Why does In silico PCR 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 In silico PCR?

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 In silico PCR.