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…in molecular biology besides PCR. Have you tried these PCR additives? How well did it work for you? What are you tips for getting GC-rich templates to work in PCR?…
…fancy way of saying “doing a PCR reaction on the amplicon of an earlier PCR reaction” or “a PCR within a PCR”. This second nested PCR will now work to…
PCR was actually one of the first lab techniques I learned as an undergrad. Despite being sometimes labeled as a pretty basic lab skill, PCR doesn’t always work as expected….
…mediator sequence results in dequenching of the UR fluorophore by release of the quencher or strand displacement. Mediator Probe PCR works with both DNA and RNA targets in real-time PCR…
…performed through genotyping PCR. What is Genotyping PCR? Genotyping PCR is used to determine the genotype of an organism (e.g., WT vs. mutant, or WT vs. transgenic). PCR primers are…
…use of a camera to detect the generation of fluorescence in ‘real-time’ as the PCR cycling was progressing. The theory worked on the simple basis that a relative amount of…
…PCR requires the addition of 7-deaza-2′-deoxyguanosine, a dGTP analog to the PCR mixture. Slow-down PCR also uses a standardized cycling protocol with varying temperatures. Generally, ramp rates are lowered, and…
…upstream (PCR #1) and downstream (PCR #2) of my gene of interest and then PCR those two fragments together for a clean deletion in the SOE process (PCR #3). The…
…witches. Well, scientific witches—by which I mean: valid, solid reasons, for why a PCR is not working! It is so frustrating to spend time and energy on a PCR, and…
…above PCR additives can improve PCR results, it is impossible to predict which additives at what concentrations are best for your PCR needs. Therefore, like always, you need to empirically…
…yet. PCR is like this sweater – it goes with almost everything and molecular biology is taking full advantage of this using it at every chance it gets. Since PCR…
…it works The first of two PCR reactions allows you to create a linear insert containing plasmid sequence at both ends. These will enable the strands of the PCR product…
…more sensitive? References 1Experts Roundtable: Real-time PCR and microbiology. In Real-time PCR in microbiology: from diagnosis to characterisation? ed: Ian M. Mackay. Caister Academic Press 2J. Clin. Microbiol. 2008 Jun;46(6):1897-900….
…incomplete extension products and it’s these guys that make PIPE PCR possible. By following certain primers design rules, the PCR process introduces short, overlapping sequences at the ends of these…
…specific PCR primers, simply add the M13 sequences to the 5′ ends. Order these from your oligo provider and your PCR is (almost) ready to sequence. PCR clean-up: You need…
I think we all have been through those my-PCR-product-didn’t-get-amplified days. Sometimes, playing around a bit more with the PCR conditions brings luck, or sometimes it doesn’t work at all. These…
…PCR Reagents and Consumables Separate Store all the PCR related tubes and vials separately from other reagents and consumables in the lab. For example, keep tubes and strips for PCR…
Routine PCR? Let’s be honest, there’s no such thing. Even with the simplest PCR reaction things can go wrong, so you need to have a good checklist of ideas for…
…can directly perform PCR using the colonies on the petri dish that the post-doc left on your bench. Some people refer to it as “Colony PCR” while others call it,…
While most may think standard Taq is the backbone of PCR, many other DNA polymerase options exist. The polymerase you use significantly impacts the efficacy of your PCR, specifically on…
…By relying on fluorescent dyes or fluorophore-tagged probes to measure the amount of amplified product, it eliminates the need for gel electrophoresis. Instead, real-time PCR instruments can measure the fluorescence…
…and pain later. For a more comprehensive guide to PCR, download our free PCR fundamentals eBook and become an expert. More reading on related topics Ten Tips for Consistent Real-time PCR…
…done by performing methylation-specific PCR on bisulfite-converted DNA. Primer design For methylation-specific PCR you include CpGs in the primer sequences. That means you have to choose whether you want to…
PCR is highly sensitive, but the downside of that very property is that it makes the technique prone to producing false-positives. In labs where PCR is a staple, like the…
…breakthrough, together with advances in oligonucleotide synthesis made PCR both cost effective and convenient and it quickly entered mainstream research. As researchers flocked to PCR, improvements to and variations on…
A wide variety of enzymes are available for PCR and RT-PCR and the optimal choice depends on a range of factors specific to your experiment. Some of these factors will…
Gene expression analysis plays a pivotal role in a wide range of studies, including biomedical analysis and diagnostics. Of all the methods available for gene expression analysis, quantitative real-time PCR (qRT-PCR)…
…Here is a brief list to help you along. Ligation Reagent Functions PCR or synthetic oligonucleotides Standard PCR reagents and oligo design techniques are used to amplify your desired fragments. …
…methods can be difficult, which is why Anchored Multiplex PCR is so great. Another strength of Multiplex PCR is its ability to enrich small and poor quantities of nucleic acid….
…assembly (PCA), or assembly PCR, is PCR’s way cooler older sibling. While both use much of the same technology and reagents, the goal of PCA is to assemble two gene-sized…
…amount of PCR product? The Mix I’ll start from the premise that you have already optimised your PCR reaction, you know the best conditions and have established the PCR cannot…
…performance of your reagents. For quantitative real time polymerase chain reaction (qRT-PCR) there may be several factors that influence technical replicates including slight differences caused by plate variability, or freeze…
…on PCR. You don’t want to lead him/her astray, but it’s hard to remember the parts that you struggled with in your early days. This article will remind you of…
…AmpliGrid, you will need an AmpliSpeed slide cycler. Two of the most commonly used options are: RT-PCR: reverse transcriptase enzyme is used during the PCR to convert RNA to complementary…
…technology is that you obtain real-time sequencing results. Today, I’m going to talk about a different NGS platform that also allows real-time sequencing, but it uses a different principle. Sounds…
…their absence can also be used to your advantage. If you are using PCR to create the insert or to amplify a linearized vector, PCR primers lacking 5’-phosphates are useful…
…Well in molecular biology, all roads lead to PCR– and away from PCR– and back to PCR again. Basically, PCR is the central technique in all of molecular biology, and…
…For all further characterizations of offspring from established mouse lines, use PCR, because it’s quick and easy. You can use a conventional end-point PCR with an agarose gel readout for…
…your PCR fail (a bit) less often? Download our free notorious PCR inhibitors poster and pin it up near your DNA engine. Or download the Bitesize Bio PCR eBook for…
You’ve tried all the usual stuff, and checked the primer sequences twice, but still can’t get that PCR fragment amplified. It’s time to enter the strange world of PCR additives….
Technique, tips, ergonomics, and ISO guidance all in one handbook
