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Genomics and Epigenetics

Codon Optimization 101

The intriguing thing about protein expression is that the combination of transfer RNAs (tRNAs) that translate the 3 letter codon into an amino acid (aa) far exceeds the number of existing amino acids (aa). If you do the math correctly, the maximum number of unique combinations using the triplet code to code for the 4…

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Maxam-Gilbert Sequencing: What Was It, and Why It Isn’t Anymore

In the mid-1970s, two methods were developed for directly sequencing DNA: the Maxam-Gilbert sequencing (or chemical sequencing) method and the Sanger chain-termination method. Indeed, in 1980, both Walter Gilbert and Frederick Sanger were awarded The Nobel Prize in Chemistry for “their contributions concerning the determination of base sequences in nucleic acids”. Actually, each got a…

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An Introduction to Nanopore Sequencing

DNA sequencing is the most powerful method to reveal genetic variations at the molecular level, leading to a better understanding of our body in physiological settings, and pathological conditions. It is the beginning of the long road towards better diagnostics and personalized medicine. Even though there have been great advances in DNA sequencing technologies there…

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A Crash Course in CRISPR-Cas9 Editing in Drosophila

CRISPR-Cas9 has become a magic tool for molecular biologists, transforming genetic engineering from a once unbelievable dream into tangible reality. Today, you can easily edit primary cells or cell lines within a few weeks with well-established protocols or others’ hands-on advice. However, no matter how many success stories you find online or hear at seminars…

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How to Design a CRISPR Experiment and Start Genome Editing

CRISPR and the CRISPR Associated system (Cas) are powerful gene editing technologies. Originally identified and characterized in bacteria, the endogenous CRISPR systems act as an RNA-based defense mechanism against invading phage DNA. CRISPR cas9 gene editing was adapted for genome editing in 2013 and has since been exploited for its ability to generate targeted double-stranded DNA…

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NGS-Based HLA Typing Delivers More Comprehensive Information

Used for matching organ transplants to donors and other applications, human leukocyte antigen (HLA) typing is rapidly shifting from older methods to NGS technologies. This is a major step forward, as more complete views of the highly polymorphic HLA genes provide a deeper understanding of how a person’s natural genetic variation might affect transplant matches…

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How to Test the Efficiency of your sgRNA

To successfully edit your genome of interest, one critical step is to test the sgRNA you have designed. Fortunately there are programs that have been developed such as CRISPRscan for zebrafish, SSC, Sequence Scan for CRISPR, or WU-CRISPR that you can use to predict the efficiency and the suitability of the sgRNA. However, the prediction…

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Meeting the BioPython

The Biopython Project is an amazing initiative that helps scientists use Python for bioinformatics – and it’s exceptionally easy to learn! You can access online services, parse (read) different file types, analyze, and do a bunch of fun stuff with your data with Biopython. The people behind the project have put in a lot of…

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Probability Theory and Molecular Barcodes

In biology, a molecular barcode is a characteristic DNA sequence used to distinguish and gather together similar items. Such a simple but powerful concept is useful in various applications. As an example, the Barcoding of Life project aims to identify specimens through the sequencing of standard gene regions, and use these as barcodes. On the other…

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Demystifying NGS: Depth Coverage and Deep Sequencing

NGS is not a three-headed monster. However, it can be a difficult concept to grasp—especially when you are getting started.  There is a lot of new terminology, and a whole new world to discover: both in the lab bench and in interpreting your results. It helps to start somewhere. So, let’s start! Depth of Coverage…

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A Primer on Checking the Methylation State of the Genome

We all know that genes encode proteins that make up a living cell. However, the level and coordination of gene expression is really the key to the success of a living cell. One way eukaryotic cells (that’s us!) control protein expression is through addition of a methyl or hydroxymethyl group on the cytosine nucleotide. This…

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Show Disparity in Gene Expression with a Heat Map

Have RNA-seq or microarray data? What possible tools can help you find your genes of interest? Is there any pattern in your expression data? I know you are totally at sea but heat maps are now commonly used to help. A heat map is a well-received approach to illustrate gene expression data.  It is an…

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Phylogenetic Tree Construction Made Easy with Blast & Mega

Your DNA sequence can be put to good use fairly easily with Blast and Mega software. These programs can help in phylogenetic tree construction. You can ask questions like what is the evolutionary relationship between a set of sequences from different species? Or how have certain microbial strains arisen? Blast As any bioscientist probably knows,…

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How to Generate High-Quality SNPs Data Sets from NGS

SNPs or single nucleotide polymorphisms are on many scientist’s wish list in experimental studies of genomic DNA sequences. Methods to detect SNPs have evolved. Now with the availability of high throughput sequencing methods, also known as next generation sequencing (NGS), SNPs can be identified in the large amounts of DNA sequence that is generated. There…

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Investigating an Expression Quantitative Trait Locus (eQTL)

Thousands upon thousands of genetic variants are now associated with every disease and trait you can possibly think of. Such traits range from cancers to blood pressure, intelligence, height, weight… and many more! This is largely because of the advent of genome-wide association studies (GWAS). However, the vast majority of genetic loci associated with these traits are…

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Variations on the ChIP-seq Theme and Challenges of Befriending Large Datasets

ChIP-seq has proved amazing. Through these new techniques, we can obtain big datasets in a matter of days, making our lives in the lab easier and more efficient. ChiP-seq combines chromatin immunoprecipitation (ChIP) assays with whole genome sequencing. This makes it possible to understand where proteins bind to DNA and epigenetic modifications. Humans are not only their…

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Introduction to Linux for High-Throughput Sequencing Analysis

So, you’ve spent time planning your high-throughput sequencing experiment. You’ve chosen how many replicates to use, deliberated about sequencing depth, and kept everything RNase-free. Now you have many gigabytes of data available. What’s next? While the first step of RNA-Seq analysis is aligning your sequencing reads to a reference genome, first you need to get…

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How to Follow up on a GWAS (Genome-Wide Association Study)

So, the genome-wide association study (GWAS) data for your disease of interest was published, and it has thrown up some very interesting associations. However, at this stage, bear in mind that this is only an association. Your project is to provide the link between the GWAS single nucleotide polymorphisms (SNP) and pathological changes. Where do…

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How Does Automated Electrophoresis Perform DNA Size Selection?

Anytime lab processes get automated by a sophisticated scientific instrument, there can be a “black box” effect, leading users to wonder what’s going on in there. For DNA electrophoresis, it’s no different. It’s easy to see what’s happening in a manual gel, but the automated gel-based DNA size selection platforms can be more mysterious. Automated…

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