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What is HRM Analysis? It is now more than a decade since the introduction of melting analysis to characterize PCR products. Melting analysis following SYBR green-based real-time PCR has become a mainstay in research laboratories worldwide for applications such as gene expression because of it’s ease of design and cost-effectiveness (i.e. no need for expensive labeled probes)….
We recently featured an article about how to streamline your cloning. But what about those days when you have too much on your plate, and need to put some things off until later? Here are a few hints on where you can pause in your cloning experiments while working on other projects: We sent a…
Northern and Southern blotting are now a thing of the past. They’ve been replaced with a faster and more quantitative technique. No longer do we wait days to know whether a gene is expressed. We can have the answer in 45 minutes! Real-time PCR is now commonly employed in almost all molecular biology laboratories to…
While almost all of you are probably familiar with the power of eBay to bring you everything from concert tickets to electronics to your very own Batmobile, you may not have realized that the world’s largest garage sale also has quite a collection of laboratory equipment. I’ve been turning to this source for equipment for…
While reading my back issues of Applied and Environmental Microbiology (AEM), I came across an interesting paper that detailed an in-depth study on the effectiveness of hand cleaners to remove Norwalk virus (NV) from intentionally contaminated hands. Yes that’s right – intentionally contaminated, and how. The study volunteers allowed a 20% stool suspension containing Norwalk virus to be…
Graduate school (PhD training) is full of roadblocks and obstacles that threaten to hinder progress, but your major professor (PI) should not be one of them. If you are frustrated with your progress and your lab environment has become unbearable, don’t throw in the towel just yet! You may need to change labs. Finding the…
We recently had a feature from Jode on everyday equipment that you can use in the lab, but what about the other way around? Do you ever take a look at what you’re doing in the lab and think, “Wow, this would really come in handy at home?” Here are a few of the things…
Reading papers on-screen is not something that everyone likes but if you can get used to it, it will help save you time and paper and make filing your literature a breeze. If you use a wide flatscreen monitor, something that is 17inch or bigger, then this tip could make your on-screen reading more pleasurable….
One of the very first things you need to do when getting set up for quantitative PCR (qPCR) is to determine the efficiency of the assay because knowing the assay efficiency is critical to accurate data interpretation. Here’s how.
I always keep an ear open for helpful tips in the lab – those little tricks that can make your experiments faster, easier and better. Here are a few tricks I’ve picked up for trimming down the time it takes to do your cloning: Restriction digestsMany digests are complete within 10 minutes of digestion at…
PCR is highly sensitive, but the downside of that very property is that it makes the technique prone to producing false-positives. In labs where PCR is a staple, like the one I work in, any false-positives are more often than not due to amplicon contamination. A broken capillary or a PCR plate left carelessly at…
The release of the iPad this week may bring the long-expected replacement of the paper-bound lab notebook by electronic notebooks one step closer. But are scientists, particularly PIs, comfortable with electronic lab notebooks? The rise of the tabletsThe concept of an electronic lab notebook isn’t anything new, and even the idea of implementing it on…
In an ideal world, every PI would be a nurturing and challenging mentor who carefully guides your project and is invested in developing your skills as a scientist. In the real world, however, that kind of leadership can be hard to find. In any case, one of the most important and useful mental steps you…
A recent article published by The Scientist called Power Couples gave advice and examples for scientist couples who have successfully balanced their life at home and in the lab. It was interesting from the perspective of how two very busy and career motivated people work together to have it all: raise a family, run a lab, and stay in love…
If you have ever attempted to load a SDS-PAGE gel only to miss the well, stab the divider, and then watch helplessly as your sample squirts off into the wrong well, then this tip is for you. The fortunate among us are able to use pre-cast gels with the wells outlined on the gel plate,…
Find out how to build a plate centrifuge using a salad spinner. Gathering the components is as complicated as it gets!
We received the following question from Bitesize Bio reader, Beheroze Sattha. It relates to a problem with absolute quantification using plasmids for standard curves. Since many people use this technique it is an interesting one question for us to explore, and it also gives us a great opportunity to cover some important tips for performing…
Presenting your work is a fantastic opportunity to get feedback on your project, demonstrate the significance of your results, and make the connections that will enhance your future career. And yet, how many incomprehensible lab meetings have we all sat through? How many seminars have you attended that left you feeling more confused than inspired?…
The humble plasmid. We now know it so well, but as little as 60 years ago the field of extra-chromosomal heredity was decidedly murky. Not only was it the subject of great debate, conflict and friction within the scientific community, it was even used as a politico-religious tool during the Cold War! The origin of…
If you’re performing DNA/RNA precipitations, you will have read Suzanne’s excellent article on which alcohol to use for precipitating your precious samples (check out some useful info in the comments for that article as well). Its publication prompted the recall of a useful tip I learned from a post-doc many years ago, one of those…
There’s something in the water, and it would love to go after your experiments. Straight out of the tap, water contains microorganisms, endotoxins, DNase and RNase, salts and other impurities that could gobble up your experiment in one bite. Of course we avoid this drama completely by using purified water from which these nasties have…
Working at a plasmid repository, I get a lot of feedback from scientists who are relieved we exist simply because that means they don’t have to request a plasmid directly from another academic lab. Either they’ve had a bad experience making requests in the past, or they really don’t know how to go about doing…
The fourth Thursday of November marks the annual tradition in the U.S. called Thanksgiving. Originally, Thanksgiving was a religious holiday that has sinced turned secular and became a national holiday in 1941. Now, for families celebrating Thanskgiving, it is a time to cook a whole lot of food and eat way too much pumpkin pie….
How often do you make errors in the lab that ruin a good experiment? Rather than flaws in experimental design, I mean errors like forgetting to add a reagent, pipetting the wrong amount, or following a protocol step wrongly. Especially early on in your career, errors like this can be a real drain on your…
TAE or TBE, which is best? Well, of course, it depends on what you want to do. Here are the pros and cons of both: But you might be better of using neither of these buffers. Despite the fact that they have been firmly established as the most popular buffers for DNA electrophoresis since their…
My PhD was a soul-less affair. It was also rock-less, jazz-less and pop-less. And all because my supervisor was of the opinion that music in the lab was a distraction that reduced concentration and our ability to do the job. “Rubbish!”, I thought, “Nothing helps you through a mindless task like splitting cells, pipetting or…
In a previous article, I listed some ways that people annoy their co-workers and many of you added some of your own pet peeves. Now I would like to discuss some ways to be the lab favorite, also known as the “golden child”. Does your lab have a “golden child”? Someone who is always perfect,…
One of the great features of PCR is its excellent sensitivity as we know. And many articles describe real-time qPCR as an added leap forward in that sensitivity – to the point where it has become a standard feature of a new assay description Indeed, I’m currently developing a new qPCR assay to replace a…
If you’re like many researchers, problems with PCR amplifying high GC DNA templates will be a major annoyance for you. Many strategies developed to overcome this issue. Betaine is the most common PCR additive used to enhance amplification of GC rich sequences because of its ability to dissolve secondary structure that blocks polymerase action. But…
You youngsters don’t know how easy you’ve got it. Kits, outsourcing and improved practices are making research easier and easier. At least in theory (who are we kidding?). In the old days things were much tougher, and many wiley old scientists bear the scars, mental and physical, of carrying out techniques that were mind numbing,…
qPCR is a technique used daily in most labs, but the first step, designing your qPCR primers, can be the biggest obstacle to even getting started. Without a good pair of primers, you can’t start asking the real questions and generating data. And sometimes the effort involved in optimizing an assay for high efficiency and sensitivity…
It’s the molecular biologist’s version of ‘I have good news. . and bad news’. The good news is that I amplified the DNA band of interest. The bad news is that I amplified these other bands as well! Oh, and this smear. What to do? Typically you might try and cut out the band of…
Hydrolysis probes, commonly also referred to as TaqmanTM is a very popular chemistry for real-time PCR. In this article I will compare hydrolysis probes with PlexorTM. But first, a quick overview of hydrolysis probes. Hydrolysis probes, an overview Hydrolysis probes are a popular detection chemistry for monitoring sequence-specific amplification in RTPCR. Just like with SYBR…
After Peter’s Introduction to Linux, those of you brave enough to accept the challenge will now have Linux installed on your machine so today I want to highlight to you some of the wonderful free software that is available for biologists using Linux. Before I start, I should mention that virtually all tools of modern…
In my last article I introduced you to the Plexor System. And from that we already know that while in reactions that user SYBR Green for detection, fluorescence increases with accumulation of PCR product, with Plexor the fluoresence decreases. Today I want to compare some other well-known features of SYBR Green chemistry and see how…
A couple of weeks ago, Nick tried to convince us that we should all be using Macs. But why would you want to use a Mac (or a PC) when you could have an operating system that: That operating system is Linux. And I think that it is high time that more bioscientists got to…
In real-time PCR, there are two primary ways to detect amplicons using fluorescent monitoring. One is intercalator-based dyes such as SYBR Green, and the other is probe-based techniques (hydrolysis or hybridization probes). All of these methods share a similar mechanism of measuring increasing fluorescence during amplification. But there is another completely different way to quantitatively…
Most people that do scientific research for a living seem to have mixed feelings about their job. Many that I know are routinely day dreaming of quitting. This contrasts with the well-established, romantic image of the dedicated scientist who loves his/her work above anything else. So what is the real story? Do scientists really like,…
So you finally got your PhD (or your masters or batchelor’s) and you are making the big switch to a small biotech company. You will probably have been hired for the specific skill set that you have built during your training, but now you have to learn to apply those skills to solve real world,…
So you have some blood stored in the -20 °C or -80 °C and you want to isolate RNA from the samples. If you wanted DNA, you would have many products to choose from. But for RNA, your choices are more limited. Obtaining RNA From Frozen Blood is Difficult Why is that? The reason is that most…
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