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Techniques

How to Transform Microalgae

What is the first image which comes to mind when you think about microalgae? Green scum that covers the surfaces of ponds? Unsightly stains on pavements and walls? Far from being a nuisance in ponds, lakes, drains and on surfaces, microalgae are fascinating microorganisms which are used to understand various biological processes. Microalgae have been…

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So You Think You Can PEMSA? A Guide to Protein Electrophoretic Mobility Shift Assay

Studying nucleic acid interactions with proteins can be accomplished using a rapid and efficient electrophoretic mobility shift assay (EMSA). This method is essentially an agarose gel electrophoresis technique that detects protein:nucleic acid interactions, as the mobility of the labeled nucleic acid will be retarded if bound to a protein (compared to unbound DNA). A lesser-known…

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Use ddRAD-seq to Study Non-Model Organisms

Reduced-representation genome sequencing has been one of the most important advances in the last several years for enabling massively parallel genotyping of organisms for which there is no reference-grade genome assembly. An implementation of the approach known as ddRAD-seq, first conceived in the Hoekstra lab at Harvard, has been widely adopted by the plant and…

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Assays: Wellular to Cellular

Out with the Old… Well-based assays have been the standard for common laboratory experiments, such as fluorescence cytometry. A researcher places a small amount of sample into a well on a plate and assays it, which produces a single data point. However, this so-called single data point is actually an average of the measurements of…

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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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How History Shaped Modern Optical Microscopes, Part One: Simple and Compound Microscopes

Having some knowledge of microscope history can be beneficial to understanding and appreciating specific configurations and components, and how they produce an optimal image. For example, practically all laboratory microscopes use achromatic objectives to partly compensate for chromatic aberration. For high-quality imaging, fluorescence microscopy, and photomicroscopy, fluorite and apochromatic objectives are used to largely overcome…

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An Introduction to Digital Images in Publications

Previously, I wrote about the different ways to use scientific illustration to communicate research. In this article, I will introduce the technical aspects of digital images and the most common image file types that scientists will encounter when preparing images for publication. Raster Versus Vector Digital images are classified based on how information is stored.…

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Reducing GC Bias in WGS: Moving Beyond PCR

WGS technologies have seen significant progress since the completion of the Human Genome Project in 2003. First-generation Sanger Sequencers were limited by lengthy run times, high expenses, and throughputs that read only tens of kilobases per run. The arrival of second-generation sequencers in the mid-2000s brought about the plummeting of sequencing costs and run times,…

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How to Improve Your WGS DNA Library

In whole genome sequencing (WGS) initiatives it is not enough to simply sequence the whole length of the genomic DNA sample just once. This is because genomes are usually very large. The human genome, for example, contains approximately 3 billion base pairs. Although sequencing accuracy for individual bases is very high, when you consider large…

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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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How to Feed Fruit Fly Larvae Small Molecules

Generally speaking, fruit flies are a great model system. Not only are they small, thus taking up very little space in the lab, but their adult lifespan is only 40-60 days, so you can track age-dependent changes without having to wait months and months. Fruit flies also display complex behaviors and more than 75% of…

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Cryofixation and Chemical Fixation for Electron Microscopy

Specimen preparation is the most important aspect of biological electron microscopy (EM), as it influences everything from the preservation of the sample itself to the kind of information that can be obtained. It is vital to define the questions you are asking of the sample before embarking on an electron microscopy project. This enables you…

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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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Emerging Model Microorganisms Take to the Stage

Estimates indicate that there may be up to 2 billion living species of organisms, each with conserved and unique biological mechanisms that are vital for survival. How do scientists understand them all? Enter model organisms. Model organisms, as the name implies, are living things which are used as representative models for understanding other organisms. They…

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Isolating Bacterial RNA from Blood

For many decades, the only way to detect sepsis – bacterial growth in blood – was isolating the bacteria and growing bacterial colonies on a special medium. This was done by first spinning down the blood, which brought the blood cells and bacteria into the pellet. The pellet was spread on a blood agar plate…

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Ultramicrotomy for Electron Microscopy

Ultramicrotomy is the process by which a sample is cut into very thin slices or “sections”, usually for imaging by transmission electron microscopy (TEM) or relatively new techniques using scanning electron microscopy (See Array tomography in three dimensional scanning electron microscopy for biology). This technique requires a bit of finesse, and this article will help…

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How (and Why) to Label Nucleic Acids

Have you ever wished you could snag individual strands of DNA or RNA with a lasso? Or look at them one by one, figuring out exactly where they are or what they are doing? Fortunately, there are techniques that exist to label nucleic acids for their visualization and purification! Nucleic acids can be labeled at…

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