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DNA / RNA Manipulation and Analysis

DIY Centrifugation-Based Purification of cfDNA

There are many reasons you may want to study circulating, cell-free DNA (cfDNA) such as non-invasive prenatal testing to generate a molecular karyotype of an unborn fetus or for use in cancer to detect, diagnose and monitor the disease. Qiagen’s QIAamp circulating nucleic acids extraction kit is consistently cited in the scientific literature as the…

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What’s The Problem With Ampicillin Selection?

Resistance to the antibiotic ampicillin is commonly used as a selection marker for plasmids in gene cloning and protein expression in E.coli and other bacteria. While it can be incredibly useful tool, there can be problems using this selection marker that you need to be aware of if if you plan on using it. This…

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A Beginner’s Guide to How Blunt-End Cloning Works

Blunt and sticky might sound dull and dirty but knowing how these different cloning methods work is important when choosing which method to use. Here we give you a guide to how blunt-end cloning works. Blunt-end cloning is the cloning of DNA fragments containing no unpaired bases at the 5’ and 3’ prime ends (i.e.…

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Antibiotics Used in Molecular Biology

Antibiotics are used in a wide range of techniques in molecular biology including molecular cloning and are important for treating pesky mycoplasma contamination in cell cultures. They can also be used to maximize your plasmid yields by reducing protein synthesis, in certain circumstances. The aim with this post is to provide an easy reference to…

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Top Tips for Troubleshooting In Vitro Transcription 

The Phobia of RNases My first experience of troubleshooting in vitro transcription came when I was synthesizing RNA In-Situ Probes for the first time. A lab mate ominously warned me that I had just returned to lab after a bout of flu and that meant I’m a walking talking factory of RNases. I went ahead…

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Faster Ligations: PEGing down the Secret

Overnight ligations are inconvenient — especially when they fail. Luckily, there’s a straightforward way to faster DNA ligations. This article highlights the secret ingredient to faster ligation reactions and offers some tips and caveats on its use. For a general overview of DNA ligations, see here and here. Buy a Quick Ligation Kit The most…

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How to Set up Your Elution Experiment

What do DNA mini preps and protein immunoprecipitation experiments have in common? They start differently, but they end with the same, critical stage – elution. But what exactly is elution, and what is the point? The Terminology First, let’s start with some basic terminology: Elution – extracting one material from another by washing with a…

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CPEC– a Quick and Inexpensive Cloning Strategy

Cloning Strategies – a Whole Lot of Options to Choose Molecular cloning has come a long way from simple restriction digestion-ligation cloning strategies to a large number of highly efficient alternatives. Broadly classified, cloning techniques can be divided as sequence dependent and sequence independent strategies. Sequence-dependent strategies are based on restriction digestion-ligation techniques or site-specific…

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Show Your Molecular TALEN(T)

Introduction Did you know that the idea of using genetic engineering to ameliorate certain human diseases was viewed as ‘science fiction’ only 10 short years ago? While cell mutagenesis studies and genetic knockout experiments were feasible before genetic engineering, they were not very reliable. Indeed, due to the random and imprecise nature of these older…

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NGS Target Enrichment Strategies

Next-generation sequencing (NGS) has ushered in a new era of understanding of both the inner workings and the function of the genome. NGS allows researchers to look at traits—including diseases—that are linked to differences or mutations in an individual’s genes. Since only about 1% of the human genome constitutes genes that code for proteins, several…

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The EMSA – Teaching an Old Dog New Tricks

Probing Nucleic Acid-Protein Interactions with EMSA The electrophoretic mobility shift assay (EMSA) is a powerful technique for detecting specific-binding of nucleic acid-protein complexes. Over the past 30 years, EMSA has been the “go to assay” to investigate the qualitative interactions between nucleic acids (DNA or RNA) and nucleic-acid binding proteins. Through the use of radio-labeled…

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