Hybridization is a common step of many, if not most, molecular biology protocols. For example: northern and Southern analysis, PCR/qPCR, cloning, in situ hybridization, array analysis, gene knockdown, and next generation sequencing (NGS).
The criteria for hybridization are based on nucleic acid strand melting. Therefore, an understanding of melting temperature (Tm) provides information on when and how the DNA or RNA strands are going to hybridize and defines the rules for hybridization. It is very important to understand this process so that you can better design and optimize the oligos for your experiments.
This is why Integrated DNA Technologies have put together this nice article so that you can understand Tm better and hopefully design better oligonucleotide based experiments.
This is probably a good time to mention IDT’s useful OligoAnalyzer online tool.
Although qPCR seems to have been around for ever, I realize that not everyone is as familiar with it as I am. It’s often easy for me to assume, when talking to my customers, that they are familiar with the terms most commonly associated with using qPCR. While I’m no expert in the use of qPCR, I’ve been involved with it in some way (i.e., on the product sales and marketing side) for close to 15 years. So, it’s easy to forget that, for some people who are just getting started in research, this stuff is mostly new to them.
Here, I’m bringing your attention to a recent DECODED article from Integrated DNA Technologies (IDT) called
I particularly recommend newbies to check out the links at the end of the article to the MIQE Guidelines and IDT’s very own qPCR Application Guide.
As you’d expect from the world’s leading manufacturer of oligonucleotides, IDT have accumulated a lot of knowledge about qPCR. You might think it is easy to just put that knowledge out there, but I know it takes a lot of effort to write articles that are genuinely useful (rather than self-serving), correct them and then spend the money to put them into print. Of course, the same information can also be made available online for free! So, I direct you to the link below (or click on the picture above) to be taken to IDT’s webpage where a Special qPCR Issue of their magazine DECODED awaits.
And of course, you should make sure to contact You Do Bio to find out how to save money and have an opportunity to try out how good IDT’s PrimeTime qPCR assays really are. I have many customers who have made the switch and are very glad they did so!
This is really great news and will mean a bit more convenience for the many Danish institutions that have chosen to use IDT as the place to purchase their DNA oligonucleotides, qPCR assays, synthetic genes etc. From this date all your transactions with IDT, from quote requests to order invoicing will be made in Danish krone.
Please contact me if you have any questions about this.
My own experience has seen the difference that automation can make to throughput in next generation sequencing laboratories. For example, I’ve been told by one customer who set up an Agilent NGS Automation system in their laboratory, that it originally took two technicians three months to process 200 samples manually. After installation of an Agilent Bravo, they increased their throughput to 500 samples per week.
Agilent are currently running a campaign on automation, with resources, videos etc. that any researchers interested in this subject will find useful.
Agricultural Biotechnology, which I guess means the application of biotechnology to agriculture, is of great interest because it is where research starts to be applied in ways that affect the food we eat, the energy we use and how we “do” agriculture. You Do Bio has quite a lot of customers who are working in this field – a couple of weeks ago I was at the Plant Biotech Denmark meeting as an exhibitor. With the general widening of the use of genomics research tools in research, it makes sense Agilent is planning their first European Animal and Plant Symposium which will be hosted in Amsterdam, February 25th and 26th 2014.
This Symposium will be over two days attracting leading scientists within the Agricultural Biology field, working in gene regulation, epigenetics, copy number/mutation detection and molecular biology using next-generation sequencing, microarrays and RT-qPCR/PCR. I think Agilent has done a great job in attracting some interesting outside speakers for this symposium. I know from experience how hard it is to get this number of speakers organized and available on those particular days of the meeting.
The meeting is open to all who work within the areas of:
|Microbiology||Animal Genomics Research|
Another focus is to investigate how such technologies and applications assist to address key concerns of the scientific community and to overcome current challenges in Agriculture Genomics Research.
|Discover the most up-to-date agenda here.|
To register for the meeting please submit your details here.
Participation is free of charge but seats are limited!
I lived in The Netherlands for 4½ years and really like Amsterdam. It is also a fairly inexpensive place to fly to and stay. I wish I was going myself!
13/09/13: eSeminar – Flexible Gene Regulation Solutions for Bacterial, Plant and Animal Sciences (Agricultural Biotech.)
Agilent are hosting a free online “eSeminar” for scientists involved in the Agricultural Biotechnology field. This will be held on Wednesday September 25th, 2013 at 1630 Central European Time (CET).
The abstract from the eSeminar reads as follows: “Today`s agricultural biologists have access to a variety of technologies at their disposal for studying genetic information. Such technologies include next-generation sequencing, microarrays, RT-qPCR/PCR and mutagenesis. By providing a complete portfolio of integrated gene regulation workflow solutions, Agilent gives you the power and tools needed to progress your science in plant, bacteria and animal genomics.
We have fully customizable content for maximum flexibility in experimental design utilizing catalog to custom microarrays and our market leading SureSelect products for non-human and model organisms.
From reproductive to crop science, Agilent has the scientific agility to shorten your time to market with customizable solutions for Agricultural Biotechnology.”
IDT have arranged a webinar with the very knowledgeable (and nice) Professor Mikael Kubista, on the subject of doing single-cell expression profiling. This is a subject that I see being of increasing interest to researchers. Perhaps it may also have future application in the fertility field, so the techniques here might end up in a clinical setting one day. Who knows?
Professor Mikael Kubista and colleagues pioneered “single-cell expression profiling” by qPCR. They have observed surprisingly large variation in the numbers of transcripts found among cells collected from seemingly homogenous tissues and even among like cells in culture. Distinct gene expression patterns can be used to distinguish cell types, subtypes, and subpopulations.
In this webinar, Kubista will describe the workflow his group uses for single-cell expression profiling and their research monitoring the activation of astrocytes in response to brain trauma in mice, stressing important experimental design aspects. He will also present intracellular expression profiling and multianalyte single-cell profiling, demonstrating the simultaneous measurements of DNA, mRNA, microRNA, lncRNA, and protein in the same cell.
There are going to be two webinar sessions. The choice of which one to go for will most likely come down to your own time zone.
Tuesday, June 25th
7:00am CDT / 2:00pm CEST / 8:00pm SGT
Tuesday, June 25th
1:00pm CDT / 8:00pm CEST / 2:00am SGT*
I think that this is a great new release by Integrated DNA Technologies (IDT, www.idtdna.com).
It’s been a long-awaited release. IDT has announced the launch of PrimeTime® qPCR Assay Plates, for high throughput qPCR analysis. Primers and probes for 5’-nuclease and intercalating dye assays (e.g., SYBR®) can now be directly ordered online in a 96 well plate format, eliminating the time-consuming transfer of primers and probes from reagent stocks, and streamlining the reaction set-up. PrimeTime qPCR Assay Plates offer an ideal solution for many applications that generate large quantities of data, such as validation of NGS and microarray data, as well as high throughput gene expression screening.
Addressing the needs of each particular qPCR assay requirement, a range of primer and/or probe concentrations are available, delivered lyophilized within a 96 deep-well plate. For additional flexibility, different dye-quencher combinations can also be ordered on the same plate, extending the range of assays that can be performed during a single run. The researcher does not even need to purchase a complete 96 well plate, just 24 assays is the minimum per plate order. Further cost savings are possible once a plate layout has been designed, since replicate plates can be generated at a lower total cost per reaction, making this format particularly suited to high throughput applications in academia and industry.
Designing and ordering your custom assays in this new plate format is rapid and intuitive with the online PrimeTime® qPCR assay tool, which allows users to easily create a master plate via simple copy and paste actions from source data. Alternatively, researchers can select from a specialized library of predesigned human, mouse, or rat assays generated by IDT’s sophisticated design engine. In addition to incorporating accurate Tm and secondary structure prediction data, this assay design engine takes into account the latest data from the NCBI RefSeq database, to avoid off- target amplification and SNPs, thus ensuring maximum assay performance.
As standard, IDT ensures the highest quality oligonucleotide synthesis, with full QC assessment by mass spectrometry, and results available free of charge on our website.
February 22, 2013 by Peter · Comments Off on 22/02/13: When You Need to Customize PCR/qPCR Primer Designs – Free Webinar
This is notification of a new webinar from IDT:
It is often necessary to select primers using specific parameters; e.g., a particular GC content or Tm, forcing the start or stop position of a primer end. However, not all primer design tools provide this flexibility. In this interactive webinar, we will demonstrate several primer customization scenarios using the free, highly popular PrimerQuest™ Design Tool, including:
• When to use default design parameters, and when to customize assay design
• How to design primers for one, two, or up to 50 sequences at a time
• How to approach specific customized design scenarios suggested by and voted on by participants of this webinar
Tuesday, February 26th – 7:00pm CST
Wednesday, February 27th – 8:00am CST (This is the best one for European-based researchers)
Wednesday, February 27th – 1:00pm CST
This webinar could be interesting to researchers who have been using IDT’s PrimerQuest, as this has been updated.
This time the webinar is from IDT and will be on Tuesday, September 25th. The nice thing about this one is the convenience: you can choose from one of two times when you would like to hear the webinar. Like all the IDT webinar series to date, I expect this one to be on YouTube shortly after completion as well.
Anyway, here are the headline and link to the information/registration page:
Presented by Professor Stephen Bustin
Those nice people at Agilent are organising an e-Seminar on MIQE, under the headline of “qPCR Assay Design and Validation.” This e-Seminar comes with the following description:
“A sensitive and specific qPCR assay is a basic prerequisite for the acquisition of reliable, reproducible and biologically relevant results.
Many published assays have been neither appropriately designed nor properly validated, leading to conclusions that are frequently wrong.
Some commercial vendors offer predesigned assays, claiming to generate optimal results for negligible user input. The quality of these assays varies considerably and it remains essential to optimise and validate those assays in laboratories.
Consequently, the ability to design assays remains a key skill for anyone aiming to use qPCR assays and the aim of this seminar is to demonstrate just how simple good assay design is.
Join this live eSeminar and learn about:
|•||How to streamline a workflow that allows researchers to design bespoke assays|
|•||Develop techniques to generate robust data|
Speaker is Stephen Bustin from Anglia Ruskin University. You can
On a related subject, do not forget to check out the excellent qPCR design tools at IDT’s Website:
I am always impressed by both the creativity and quality that goes into BioCision’s products. I guess the company motto could be “death to ice,” or similar! This is particularly apt with the release of BioCision’s CoolBox XT Workstations. In summary, the CooBox XT Workstations provide over 16 hours of ice-free cooling for your samples without electricity or batteries.
If you are serious about reducing variability in the way your samples are handled at the lab bench, I believe that it is more than worth your time checking out BioCision’s product line.
There is a CoolBox XT Workstation promotion that you can also take advantage of here.
A very quick video explains the concept:
My advice is to use IDT’s PrimerQuest.
I have been working with some customers in Denmark to help get them started on this, because they are studying organisms that are not included in the (pretty big) model organism defaults to be found in IDT’s other design tools (SciTools).
PrimerQuest is pretty easy to use and I’ve posted a very short “how to” guide here. The only time-consuming part of using this tool is the BLAST you need to do on the primers/probes, but come on, if you’re in research isn’t that your job?
Following the successful qPCR assay design seminars held at the start of March 2012, a number of customers took up the opportunity to try out free qPCR assays. I recently got some nice feedback from one such customer. Always good to know that sometimes people can be helped.
This from a customer at Skejby Hospital in Aarhus:
“ I have tried the IDT ZEN probes and they are as I expected – much better than the ones we had, and our technician have ordered the real ones to use in our diagnostic assay “right away”.
It was very convincing with much lower background. Everybody was convinced by first look at the results!!!
I am happy that I got the opportunity at the right time :-)”
It is great to be able to find further information on IDT available on the internet. I think that the following will be interesting to a lot of customers who are currently using IDT: A video of the IDT European facility in Leuven, Belgium. The US sites will have a lot of similarities of course. Have you ever wondered what a world-class oligonucleotide synthesis facility looks like? Wonder no more…
Agilent are running a bunch of promotions on genomics-related product this Autumn (actually it’s turning out to be the nicest time of year, given the wet Summer). They are summarized here:
- Bioreagents: Buy 3 Stratagene products, get the cheapest one for 1 DKK
- Brilliant III: Buy 1 kit of Brilliant III qPCR reagents, get a second one for free
- Genespring & microarrays: Get 50% off your Genespring data analysis software annual licence when you order > DKK 112K of genomics products
- SureSelect: Buy 1 SureSelect target enrichment kit and get a second one for free (valid for 5 to 25 reaction kits)
- SureCycler bundles: Combine our SureCycler 8800 PCR instrument with our Mx 3005P real-time PCR instrument or with our microchip electrophoresis system (Bioanalyzer 2100 or Lab 901 TapeStation) and get 30% off your bundle
You can also go to the Official Agilent Autumn Promotion page.
Save time by contacting me directly if you are interested.
IDT have posted a short article on their website about using their PrimeTime qPCR Assays in multiplex assays. The PrimeTime assays themselves allow researchers to change the primer:probe ratio and use well-separated fluorescent dyes on the probes. Read the article to get further insight.
Just a short post to inform researchers in Denmark about the upcoming MIQE Events in Copenhagen and Aarhus.
I am very lucky in being able to get Raza Ahmed from Agilent’s genomics team over for a few days. Raza has tons of experience in qPCR, having worked for Stratagene before they became part of Agilent. Raza has agreed to give an introduction to MIQE (read more on the Events page) that should serve to educate and inform researchers about the guidelines. There will be plenty of time to ask Raza about qPCR subjects not related to MIQE as well.
Please come along, say “hej”, listen and learn and enjoy a coffee.
I am pleased to inform you (well, if you are a researcher in Denmark at least) of Agilent’s Summer promotion on genomics products. The promotion is valid from now(28 June 2011) until August 31st 2011. Headlines below, with a bit more detailed information beneath each heading
Update 12 July: Agilent have a Summer Promotion webpage now live.
Purchase the New SureCycle PCR Instrument Together with the Mx3005P qPCR Instrument and get 30% off the Bundle
This is possibly the best offer of the whole promotion. The SureCycler is the hottest new instrument from Agilent, very cool (connect to it from a mobile device) with great specs. The Mx3005P is a well-proven and robust qPCR instrument that won’t let you down.
Purchase the Bioanalyzer with Agilent SureCycler 8800 or with >5000€ of Bioreagents and get 30% off the Total Bundle
Hmm. On second thoughts, perhaps this the the best of the Summer Promos. It really depends on what you are after. Of course, Bioreagents can mean a lot of things: PCR enzymes/kits, nucleic acid sample prep and qPCR reagents would be the most obvious. The combination of the Bioanalyzer with qPCR reagents sounds like a good combination to me, given that the Bioanalyzer is great for doing QC on RNA.
Get 50% off Absolutely RNA extraction kit when you purchase Bioanalyzer reagents
For those of you who are reguarly ordering Bioanalyzer kits (and that means lots of you), this is a great opportunity to try out one of Agilent’s best nucleic acid purification lines. The Absolutely RNA extraction kits come in a number of size options (mini-, micro- and nano-prep) and there are kits for miRNA and FFPE samples too. This is really a great excuse for you to try out these kits, especially if you haven’t had a lot of success with what you are currently using.
10,000€ discount on your genomics consumables order when you purchase an Agilent Bravo Automated Liquid Handling Platform
OK, so not everyone has the money available for an Agilent Bravo. However, if you are in the market for a small-footprint, very fast and accurate pipetting workstation, you really have to consider the Bravo. You will save many many days of work by having a Bravo in your lab, not to mention saving your pipetting thumb!
10,000€ (about DKK 75K) will get you a lot of genomics consumables.
Genespring and exon arrays: buy 5000€ of exon arrays and get 25% off your Genespring annual license, buy 10k€ of exon arrays and get 50% off the annual license.
Yes, it is that simple. Purchase Agilent’s high-quality exon arrays and get a big saving on the GeneSpring license (which you can then use to analyse all that lovely data and put into a biological context).
So that is the summary of Agilent’s Summer Promotion on genomics products. It looks pretty good to me and will mean that you come back from your Summer holiday with new gear to get into your research again after the break.