DNA / RNA Manipulation and Analysis
Lab Basics: How Alkaline Lysis Works
Alkaline lysis for plasmid isolation? That’s like the ABCs in a molecular biology lab. Read this detailed article to understand the process behind this common technique.
Read More A Complete Guide to How Nucleic Acid Extraction Kits Work
Nucleic acid extraction kits are routine in today’s molecular biology labs. Read on to know about what is inside these black boxes of wonder and how you can get the best results for your preps!
Read More How to Identify Supercoils, Nicks and Circles in DNA Plasmid Preps
Why do you get three bands when running uncut plasmid DNA on agarose gels. Discover the answer and how it can help improve your DNA plasmid preps.
Read More Which Type of Ethanol Should I Use?
Want to know more about ethanol grades commonly used in the lab? We help you make sense of your flammables cabinet with our rundown of the ethanol grades typically used in molecular biology, as well as some important rules for how to use them correctly.
Read More 5 Ways to Clean Up A DNA Sample
Are you struggling with your DNA clean-up? Then check out our top five methods so you can pick the best option for your experiments.
Read More Ethanol Precipitation of DNA and RNA: How it Works
Using ethanol precipitation to isolate or concentrate your nucleic acids? Find out how this routinely used technique works, and get tips to produce the best results.
Read More How to Get a Picture-Perfect Agarose Gel
Check out our agarose gel hacks for troubleshooting your blurry or uneven bands, and tips for getting picture-perfect agarose gel images every time.
Read More Preparing Metaphase Spreads: The Breakdown on Broken-Down Cells
A quick look at the first steps of metaphase spreads – the break down on breaking down your cells and the factors to keep in mind.
Read More Five Ways to Modify Your siRNA for Improved RNAi
Want to increase siRNA stability and efficiency? Read on to learn five chemical modifications that will help.
Read More 3′ mRNA-Seq is Transforming Gene Expression Analysis
3′ mRNA-seq is a NGS method to profile gene expression that is sensitive and straightforward but won’t blow your budget.
Read More Isolating Nucleic Acids From Arabidopsis Seeds (And Other Tough Seeds)
While DNA and RNA extraction is a pretty common technique, it isn’t always the easiest. Read on to learn some top tips on how to successfully extract nucleic acids from even the toughest samples, like Arabidopsis seeds.
Read More How to Isolate DNA from Mitochondria: An Energetic Guide
Mitochondrial DNA isolation can be time-consuming and laborious. Find out how to minimise the time needed for its extraction, while ensuring fantastic results.
Read More miRNA Expression: How to Choose the Right Profiling Platform
When it comes to profiling miRNAs there are lots of platforms available. We discuss the pros and cons of various miRNA profiling methods to help you choose the right one for your needs.
Read More T4 DNA Ligase: The Only Ligase You’ll Ever Need?
T4 DNA ligase is the swiss army knife of ligases, but it can’t always do it all. Find out what it’s good at and the alternatives available for the things it struggles with.
Read More Tips on Restriction Digests: Designing a Restriction Screen
Restriction digests are the cornerstone of many techniques. Here are some great tips on setting up and troubleshooting them.
Read More Ye Olde Antibiotic Plates: Stability of Antibiotic Agar Plates
After a late night transformation you realise you have forgotten to make any plates. Should you use the old stash of amp plates you found in the back of the cold room?
Read More 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…
Read More 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…
Read More 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.…
Read More 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…
Read More 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…
Read More 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…
Read More 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…
Read More 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…
Read More 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…
Read More 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…
Read More 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…
Read More 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…
Read More 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…
Read More The Beginner’s Guide to Reading Plasmid Maps
Very often plasmid maps, especially historical ones that are hand-drawn by a long-forgotten PhD student, are a puzzle. What are these arrows and boxes? Where do I start? Don’t worry, we have a crash course introduction into deciphering plasmid maps. Familiarizing Yourself with Your Plasmid of Interest Let’s start with a classic plasmid: pBR3221. It…
Read More RNAseq Library Preparation: From Cells to cDNA
RNAseq libraries, also called whole transcriptome shotgun sequencing libraries, provide a snapshot of cellular processes. This allows the researcher to gain information regarding changes in transcriptome in response to environmental changes, during disease, or after a drug application. RNAseq libraries also allow for the detection of mRNA splicing variants and SNPs. RNAseq libraries have virtually…
Read More Picking the Right DNA Isolation Kit for Your Application
If you plan to work with purified DNA in the lab, it’s likely that you will use a commercial DNA extraction kit to isolate and purify your DNA of interest. With so many types of kits available, it can be a major challenge to choose the best one to use when working with an unfamiliar…
Read More Are You In(to) Situ? – Putting Together Your First RNAscope® Assay
You are thinking of trying out RNAscope®. After all, RNAscope® holds promise for increasing the sensitivity and specificity of your in situ hybridization. Yet, getting started can be a little overwhelming with the numerous kits and reagents available in the RNAscope product line. Here’s an overview of your options to help you navigate to the…
Read More Multiplex Ligation-dependent Probe Amplification (MLPA)
Multiplex ligation-dependent probe amplification (MLPA) is a molecular technique developed by MRC-Holland back in 2002. In a nutshell, MLPA is a sensitive technique that allows quantification of nucleic acid sequences, quickly and efficiently. It is performed in many laboratories worldwide, and can be applied to detect copy number changes (like deletions or duplications) of a…
Read More RNA Strandedness: A Road Travelled In Both Directions
For most molecular biology purposes, DNA is thought of as a string of nucleotides running from 3’ to 5’, and the corresponding mRNA sequence is complementary to this DNA string. However, visualizing this quirky DNA structure for what it is – two antiparallel strands joined together – it quite important for many applications, such as…
Read More The Next Big Thing: Alternative Polyadenylation
What Is Alternative Polyadenylation? Processing of mRNA and its regulation plays a fundamental role in gene expression. As science progresses, alternative polyadenylation takes center stage in the undercurrents of gene expression. 1,2 Polyadenylation is part of the pre-mRNA maturation process and involves polyadenylation of the 3’ end of the emerging RNA. This process happens to…
Read More The Importance of Non-coding RNAs
What Are Non-Coding RNAs? What was once considered “junk” may end up being the most important part of our genome. Non-coding RNA (ncRNA) is RNA that is transcribed from DNA but diverts from the “central dogma” because it does not code for proteins. NcRNAs are ubiquitous in eukaryotes: while 90 percent of eukaryotic genomes are…
Read More Protocols for Cloning Without Restriction Enzymes or Ligases
There are many cloning methods that do not require restriction enzymes or ligases. Read below to learn about how to achieve seamless cloning results via Topoisomerase cloning, SLIC, and Gibson. Method #1: Topoisomerase Technology Topoisomerase technology requires no special primers and no ligases – it is as easy as cloning comes. This technology is based…
Read More Cloning Methods: 5 Different Ways to Assemble
Over the past few decades molecular biologists have developed procedures to simplify and standardize cloning processes, allowing vast arrays of artificial DNA structures to be more easily assembled. Are you familiar with all the cloning options out there? Let’s look at five different cloning methods you can use to get your construct. At the end…
Read More Ligation Independent Cloning Primer Design
Ligation independent cloning (LIC) is an easy and effective method to ensure successful cloning, all without the need for ligation. As easy as the technique is, designing primers can be a bit tricky. In this article, we will present a quick overview on primer design for ligation independent cloning.
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