Olwen Reina's Profile

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Seamless Ligation Cloning Extract (SLiCE) Explored and Explained

Traditionally, if you’re hoping to clone a DNA/RNA fragment (or insert) into a vector, such as a BAC you would need: Expensive exonucleases, called restriction enzymes: pacman-like enzymes that chomp at specific sequences in your destination vector or fragments to be inserted (often just “inserts”). Sequence homologies between your inserts and your destination vector, called […]

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PCR on a shoestring: Build Your Own PCR Machine

Maybe you’re a teacher hoping to do something different, maybe you’re a student trying to drag out their funding, or maybe you just want to build something really cool. Did you know that PCR used to be done by hand? Imagine three water baths with thermometers, a timer going off every few minutes, and a […]

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In PCR, qPCR and qRT-PCR 6th of May, 2015
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Polymerase Incomplete Primer Extension (PIPE) Cloning Method

PIPE PCR is a ligase-independent, restriction enzyme-free cloning strategy like SLIC (link to my SLIC article), SLiCE and CPEC. The PIPE method eliminates sequence constraints and reduces cloning and site mutagenesis to a single PCR step followed by product treatment. It is fast, cost-effective and highly efficient. The key step is designing the primers; one […]

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In Lab Statistics & Math 4th of May, 2015
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How to Be Greener – The Environmentally Friendly Guide to PCR

Science is an expensive business and those who use high energy-demanding techniques may not even realize just how expensive they are. The Cost of PCR Let’s looks at PCR. You need to pay for the machine, all the ingredients including expensive enzymes, a freezer and a fridge for your ingredients, tubes and caps, not to […]

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In PCR, qPCR and qRT-PCR 19th of March, 2015
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The 10 Unspoken Rules of Working in a Lab

There are so many unspoken rules to working in a lab! It’s unnerving what will cause frayed nerves to snap, people not to trust you and a good relationship to turn sour. Here are some of the rules I’ve learned. Feel free to add more in the comments section below. 1. Thou Shalt Not Touch […]

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In Basic Lab Skills & Know-how 9th of February, 2015
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Finding Nemo: Understanding Single Cell Isolation and PCR Amplification

Every protocol for single cell PCR can be broken down into two steps. In the first step, the cells are isolated by micromanipulation, laser capture microdissection, flow cytometry, or by direct micropipetting. Next, the genetic material is processed by PCR to amplify your sequence of interest. Here, we’ll go through the different options for isolating […]

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In PCR, qPCR and qRT-PCR 5th of February, 2015
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Six Ways to Measure T Cell Responses

T cells can be problematic to characterise because they have a wide variety of subtypes and because of the technical difficulties of studying the membrane-bound T cell receptor, but there are situations where you want to be able to do this such as analysing the degree to which immunological memory has been induced to measuring […]

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In Cells and Model Organisms 21st of January, 2015
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Letting Go of Your Ligase: Transfer PCR

When I buy a new sweater, I love finding out that it goes with several pairs of pants, the scarf that’s an awkward color and the earrings I haven’t worn yet. PCR is like this sweater –  it goes with almost everything and molecular biology is taking full advantage of this using it at every […]

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Say Goodbye to Restriction Enzymes and Ligases: An Introduction to Sequence and Ligase Independent Cloning (SLIC)

SLIC, or sequence and ligase independent cloning, was developed by Li in 2007 and published in Nature Methods. What makes it a Nature Methods worthy protocol? Unlike other forms of cloning, SLIC does not require restriction enzymes or a ligase! Seriously! Don’t believe me? Why not have a go for yourself? I’ve detailed the main steps below to get you started. How it works To […]

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In DNA / RNA Manipulation and Analysis 21st of November, 2014
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Thermal Asymmetric Interlaced PCR (TAIL-PCR)

What do bunnies, coins and PCR have in common? They all have tails! (ha ha!) Thermal asymmetric interlaced PCR or TAIL-PCR is used to sequence and analyse unknown DNA fragments that are adjacent to known sequences. Think of it as being rather like networking. You know you want to get to know someone so you […]

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In PCR, qPCR and qRT-PCR 26th of October, 2014