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There is a growing trend in using microalgae as the expression system for heterologous proteins. However, I find most protocols dealing with microalgae available online are not that great or informative! So I would like to share my experience in using Chlamydomonas reinhardtii as the model microorganism. Before you start expressing your protein, you need…
Two years ago, all I knew was third BASE in my baseball field and the cutter ball from the pitcher. Now, I know a lot more about lab-based BASES and cutters: REBASE and NEBcutter. While they sound like baseball terms, REBASE and NEBcutter are tools for working with restriction enzymes. Read on to find out…
Researchers have always been in search of model organisms that can be used to study and explore biological phenomena to make discoveries that can be extrapolated to more complex higher organisms like humans. Of the various model organisms developed and used, starting from the ubiquitous E. coli and S. cerevisiae to the humble D. melanogaster…
Through many trials, and lots of error, I learned that there are many considerations for mass spectrometry that might not be obvious to you as a molecular biologist. Common contaminants, even in small quantities, can mask important peaks in your mass spec data and have a huge impact on the final results.
In my last article, I talked about the basic protocols and experiments conducted in the process of converting algae into biofuel. Our ability to culture algae has efficiently improved over the years. Continuous improvisation in basic techniques has helped us to understand the growth limiting step of algae culture. In this article, I will discuss…
An academic lab is a unique working environment. Lab members are expected to take responsibility for their own research projects and perform the work quickly and efficiently. However, unlike an industrial or corporate setting, there are often no clearly defined management structures. This means that when it comes to communal equipment, reagents and resources, individual…
So you want to work with mouse B cells? Primary murine B cells are a difficult, yet fascinating system to work with and can help deepen your understanding of an immunological system. You can study many things with primary B cells, including: immune activation antibody production cell-cell interactions between immune cells and immune phenotype These…
Put in the time now to optimize protein solubility so you don’t have to suffer later! With this in mind, here are some considerations and tips for selecting the optimal conditions for recombinant protein expression and purification.
Routine PCR? Let’s be honest, there’s no such thing. Even with the simplest PCR reaction things can go wrong, so you need to have a good checklist of ideas for PCR troubleshooting and rectifying the problem. Today I have brainstormed all of the ways I can think of to approach problems with standard PCR reactions….
The whole TLC technique sounds easy to do, but it can be difficult and tricky during interpretation or give unexpected results, especially when working with biomolecules. For this reason, it is important to be familiar with troubleshooting thin layer chromatography. Some of the common problems faced during TLC and their solutions are listed below: Solvent…
There’s piloting a brand new technique for the first time. Then, there’s jumping through hoops trying to get an established lab technique to work. The former, in contrast to the latter, is expected to be fraught with hardships. Yet troubleshooting an old lab technique that isn’t working anymore, is frustrating at a whole new level….
To isolate your protein, you are going to need to tag it with something that will allow you to fish it out of the bacterial protein soup. This is where transition metals can help you out!
Like most other chromatographic techniques, thin layer chromatography (TLC) separates out individual compounds from a mixture depending upon the polarity of each compound. The solvent system travels up a silica plate by capillary action and passes over the sample that you spot onto the plate. As the solvent travels up, it moves the compounds present…
You’ve tried all the usual stuff, and checked the primer sequences twice, but still can’t get that PCR fragment amplified. It’s time to enter the strange world of PCR additives. Over the years a variety of additives have been shown to enhance PCR reactions in certain situations. Here is a summary of some of the…
RNA purification may be a common procedure in molecular biology but it is by far the one that people fear most. Why? Dreaded RNase. It’s everywhere… all over your bench and pipettes, and floating in the air, waiting for the chance to creep into your prep, shred your RNA into nucleotides, and ruin a day’s…
Want to save money on one of the most expensive steps of your Western blots? Then, use less antibody for significant cost savings!
The ability for DNA polymerase to copy a long stretch of DNA is becoming increasingly important. Why? It has to do with the advances in our sequencing technologies. Our next generation sequencing (NGS) technology requires the DNA polymerase to copy a long stretch of DNA (sometimes up to 50kb) as NGS is churning out genetic…
What do you use if antibodies are too large for super resolution microscopy? Aptamers. These are small affinity reagents (~ 2 nm!) that interact with their target in the same way as an antibody, but without the hefty backbone attached.
Say you just joined a lab and have been assigned your very own project to work on. As part of your new responsibilities, you have to breed and maintain the mutant (or transgenic) mouse line which you will be using for your experiments. An integral part of mouse genetics experiments is determining the genotype of…
Hope you had a heavy breakfast, because the first qPCR is going to take time. Did you remember to fill your coffee cup? There are a lot of intricacies in qPCR that will need your neural networks to be brisk. How qPCR differs from traditional PCR Unlike traditional PCR, qPCR measures the amplification of DNA…
In the sci-fi novel Terminal World by Alistair Reynolds, a planet consists of zones with defined characteristics of matter interactions on a subatomic level. These conditions permit different levels of technology sophistication in various zones. For example, in the “Steamville zone” nothing more complicated than steam engines works – electronic schemes fuse irreversibly. Something like…
Here’s a tip that you may find useful if you are expressing proteins in E.coli using a lac promoter-based expression system, e.g. pET, in LB medium (L-broth). Lac expression systems are typically induced in the lab using IPTG (isopropyl-beta-D-thiogalacto- pyranoside), which is a non- hydrolyzable analog of lactose, the natural inducer of the lac operon….
You have a new plasmid, now what do you do? You are excited to go further with your project. But before you can move on, you have to confirm the presence of your insert as well as the sequence and orientation of the insert. Is the insert the right size? Most people use restriction enzymes…
When I began a master’s program in 2008, the lab task I hated more than anything was running agarose gels for DNA. Something so simple, ubiquitous, and necessary gobbled up more hours in the lab than I care to remember. Even though we added the DNA stain directly to the molten agarose and didn’t have…
When you first start out using a microscope, you might only adjust the eye pieces, objectives, and the focus controls. However, you shouldn’t overlook the microscope condensers as they are an important part of the whole optical system of a microscope.
The more experienced hands in your lab know that molecular biology is rarely just a journey from A to B. As a result, I’ve constructed this short workflow as an introduction to genomic molecular techniques.
Most eukaryotic proteins exist as several isoforms, differing in posttranslational modifications, which allows them to perform slightly different functions or the same function under slightly different conditions. A common posttranslational modification of proteins is glycosylation.
As biochemists, we routinely run SDS-PAGE to analyze our proteins. Imagine the time and effort you are going to save when you can run every gel to perfection.
One of the most crucial steps in any cloning procedure is the preparation of the vector. Get it wrong and your chances of success will be drastically reduced. The overall aim for a good vector preparation is to obtain a fairly concentrated stock of undamaged, fully digested plasmid DNA that is free from contaminants. Missing…
Do you know about algae and their potential in today’s world? Do you know how to work with algae? Algae are becoming increasingly important in the research world.
Producing lentiviral or retroviral vectors is theoretically fairly straightforward. However, anyone new to viral vector work is usually confronted with vast amounts of confusing information. It seems that anyone who has ever made a lentivirus has their own protocols and is adamant that their method is the best one to follow. In reality, there are…
Pat yourself on the back, you saw a post with the word “coding” in the title and you didn’t freak out or glaze over. That’s the first step. Coding seems to have such a stigma attached to it; people tend to think that it’s incomprehensible nonsense that they could never learn, and that it has…
Some tissues are tricky to work with. This truth was lost on me in the early years of grad school because I worked with liver samples. If you’re extracting RNA from liver samples, you’re likely not losing sleep over your massive RNA yields. But for the folks doing RNA extractions with less willing donors, such…
In Part 1 of this article, I introduced you to using code for basic image manipulation in ImageJ and working with the command recorder to expand your coding vocabulary. I covered how to make a simple macro, how to edit it and then save it to be run again another time. If you skipped the…
You had been happily measuring your DNA samples with your NanoDrop for a while, until… you noticed that your results are not reproducible *gulp*.
Read on – here are 10 points that will help you troubleshoot the issue.
Here are some ideas to make your liquid handling robot perform at its best.
You open the incubator in the morning and to your dismay there are a hundred glorious colonies… on your vector-only control plate. While there are a number of potential causes, I’ll highlight a few of the more likely culprits and their solutions.
Sometimes, you have to work with radioactive materials in the lab. If you happen to be one of those lucky individuals, it is important to take extra precautions in your work to ensure your safety, and good results! Here are my tips to make sure you radiate confidence doing your experiments. Know Your Element It…
I think we all have been through those my-PCR-product-didn’t-get-amplified days. Sometimes, playing around a bit more with the PCR conditions brings luck, or sometimes it doesn’t work at all. These days we have access to many different types of DNA polymerases, ultrapure and buffered nucleoside triphosphates, and other necessary starting materials in convenient concentrations; but…
The ELISA (enzyme-linked immunosorbent assay) is arguably one of the most important and versatile tools in the toolbox of molecular biologists, biochemists and diagnosticians across the world. Defined by its simplicity and speed, the assay is easy to learn and perform in as few as five steps. But with so few variables to manipulate, an…
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