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Archive for April, 2009|Monthly archive page

What you always wanted to know about viruses

In animation, Cool stuff, Swine flu, virus on April 30, 2009 at 10:02 am

Considering that swine flu is here everywhere and many became interested in viruses, I compiled here a few videos that might give you an update about viruses, virology, vaccine production and how a vaccine acts. Feedback and questions are welcome at

balintblaszlo(at)labtutorials(dot)org

Regards,

Balint.

Where are the swine flu cases?

See actual status here

or check it in GoogleMaps here:


An overview of the status by 20090430:

healthmap200904301

How does influenza spreads and what happens with it in our body?

The life of a virus

The structure of viruses

Vaccine production(description and images).

Vaccine production at Sanofi (video).

How does the vaccine act?

Injection of the vaccine

More about pandemics, and vaccination here.

How microarrays can be used for rapid characterization of viruses: here.

Stay tuned!

Bird flu solved, swine flu arrived!

In Cool stuff, News, virus on April 29, 2009 at 4:12 pm

The bird flu solved by vaccines, now comes the swine flu.

Pandemics shaped the human population in past and probably in future. One of the most destructive influenza pandemics in the last century was the Spanish Flu which caused the death of 25-40 million people. More info about the pandemics here. The very last thread to cause a pandemic is the virus called A/H1N1, or swine flu, or Mexican flu.

Rapid diagnostic tests will be probably be PCR based. A very interesting microarray based virus characterization tool is described here.

Influenza vaccines are produced by quite a lot of manufacturers in the world. You can have a list of the manufacturers here.

A small Hungarian company is a word leader in vaccine development. Omninvest was one of the first who produced the anti H5N1 vaccine. It is good to draw the conclusions of the Bird Flu vaccine development, a detailed overview can be found here.

Today Omninvest announced that they made all the preparatory steps needed to start to develop a Swine Flu vaccine. Once the sample virus arrives they can start the development of inactivated viruses that can be used for the development of the Swine flu vaccine. While US based CDC researcher Ruben Donis announced that the “reference strain” would be available at the beginning of May, the Hungarian news agency MTI released the information that samples that could be used for further development migh arrive in days.

For more details about the life of a virus and vaccination check this.

Interview with Howard Wolinsky the US correspondent of EMBO Reports

In Interview on April 29, 2009 at 10:08 am

Dear All,

In this changing world we all feel the pressure to find novel ways to increase our efficiency and impact. Recently I just came in contact with Howard Wolinsky a freelancer scientific and medical journalist. The way how Howard changed the pace of his work seems to reflect the “flat world” paradigm. We will discuss about journalism, being a freelancer, scientific journalism, and the impact of web based networking via Facebook, LinkedIn, Twitter and so on.

So let us have a short interview with Howard Wolinsky:

—————————————————————————————-

LTB:  After working so many years as an employee, you decided to become a freelancer. What was your strongest motivation to make this step?

HW: I started out as a newspaper reporter in a small town, Kankakee, Ill., USA, 1970, covering mental health (there were two large mental institutions in town) and local government.

But I had my eyes on bigger things. I wanted to be a health/medical reporter for a major daily.

My colleagues often described me as “entrepreneurial.” That meant, I freelanced and they didn’t for the most part.

I started to freelance articles to the Associated Press and United Press International. I did a story for the Chicago Daily News–an historic newspaper where Carl Sandburg, the poet, and Ben Hecht, the playwright, who authored “Front Page,” had worked–on a mental health topic. I was offered another assignment.

But the Kankakee Journal nearly fired me for writing for a “competitor.”

But I kept freelancing throughout the years. Some years, as I moved on to other papers, including St. Petersburg (Fla.) Times, Florida Today (then in Cocoa, Fla.) and Chicago Sun-Times—my freelance wages matched or exceeded my base pay.

I considered leaving to be a full-time freelancer a couple times. But one of the problems in the US is health insurance. One time, on the verge of freelancing full-time, I contracted pneumonia and was out for a couple weeks. My pay at the Sun-Times continued. But had I only had freelance, I would have had financial worries. No work, no pay as a freelance.

In the intervening years, my wife returned to work and could offer us insurance coverage.

As the economics of the newspaper business deteriorated in general and at the Chicago Sun-Times in particular, I had an opportunity to take a buy-out after nearly 27 years on the job as a health and tech reporter. As I described in an article in GaperBlock.com, I had a flash of insight on how I could take a year’s pay as “seed money” to start my own freelance business.

I left on a Friday in January 2008. The next Monday I had my first assignment.

But in fact, I unconsciously been preparing for this move since 1970.

A friend once advised me that I shouldn’t just have one job but multiple ones. That’s what I did last year. In addition to freelance writing, I diversified. I went in new directions. I networked.

I became the US blogger for Skype. I taught at Medill Graduate School of Journalism at Northwestern University. I wrote for university/medical school magazines, such as Cleveland Clinic, Northwestern Memorial Hospital, Roswell Park Cancer Center, Howard Hughes Medical Institution, University of Illinois at Chicago. I wrote for BusinessWeek, more general, non-science stories. (Check out my story in BW on David Axelrod, President Obama’s advisor.) I was invited to contribute to Huffington Post, which doesn’t pay (yet) but gave me a chance to stretch and write in new areas. I wrote for my former competitor, Chicago Tribune. I wrote some articles for Venture Capital Journal.   I did some consulting. I started doing more for my friends at EMBO Reports. I did a number of stories for WebVet.com. On and on.

And as busy as I was, I spent two months traveling. My wife and I celebrated her birthday and the Inca New Year at a yoga retreat in Peru (we were remarried by a shaman at Machu Picchu). The BBC paid my way to UK to appear on the Coast program. We went to Paris on the train for a day. Took a couple trips to Northern California and one to Florida.

LTB: You are doing plenty of different type of activities: you write books, you are a blogger for Skype, US correspondent for EMBO Reports, you are teaching and this is only the tip of the iceberg. How do you manage to keep your focus? What is your secret of productivity?

HW: Some days, I do feel that I have taken on too much. I am a fast writer, which helps. I also have turned away work if I didn’t find it interesting, but I always tried to steer the work to a friend. It’s important to network. (One of the things I did everyday after I left my full-time job was spend an hour a day building my network at LinkedIn, Facebook and now Twitter. Jobs have come my way because people have found me in LinkedIn and Facebook–even though I can be readily found through a Google search.)

LTB: You are located in Chicago but a substantial part of your activity is targeting a much wider audience. Do you see a trend in this? Is the world really becoming flat? From practical point of view do you think people need a strong local connection network and mix this with a web based business or you see that targeting people trough the web could be enough in itself?

HW: I always told my kids to think globally because that’s where the work will be. I also told them to become fluent in another language. (I am working at that now. I am taking Spanish via Skype with a tutor in Guatemala.)

The Web offers the tools to reach out and be a world citizen/worker. I ran across my phone bill from a few years ago, I cut US $2k off by using Skype. Before I became a blogger for Skype, I wrote an article for the ScienceWriters network urging my colleagues to use VOIP to expand their network of sources and also clients. Everyday now, some 350,000 people sign up for Skype. Over 400 million already have. It’s amazing–and I’m not just saying that because I am a blogger for Skype.

Seemingly silly social media, such as Twitter, are and will become increasingly important. Some people I run into seem to feel they already take up too much time doing e-mail. They reject doing Facebook and Twitter. I even talk to young people who feel that way. Some see it as an invasion of privacy. They feel teched out. Get over it.  I say give it a try. Facebook and Twitter can be about more than what you had for breakfast.

LTB: As a freelancer, can you plan your work for longer time periods (I mean more than 3 to 6 months)?

HW: I do the best I can to structure my time so I can count on a revenue stream. So far, I can rely on Skype and Medill as a base. Everything else is gravy. It’s hard to know what’s coming. I am helping develop a new magazine now, but this won’t last. I have a line on a potentially lucrative consulting job in the fall, but I can’t count on it. Clients come and go.

LTB:  What was the most important for you to have a successful transition from employee to freelancer?

HW: In the US, make sure you have health insurance. I had the advantage of working in the field for decades. It would be different if I were totally starting out. Networking is vital. Some writers I know are great networking to cover a beat, but may don’t have a clue on networking in the writing business world. They’re going to have to learn.

When I decided I was leaving my job, I wrote down every e-mail address I had and sent out notes announcing my departure. The networking helped. Contacts came up with leads that turned into jobs.

I had belong to the National Association of Science Writers for 25 years. Never went to a meeting. Until last year. I walked away from my first meeting with a lead on the teaching position at Medill. I have joined several other groups and have attended meetings. You never know when one thing leads to another.

Help others find work. They may return the favor.

Use LinkedIn, Facebook and Twitter.

LTB: With your experience now, at what stage of a career would you recommend younger people to start their preparation to become a freelancer?

HW: Maybe we’ll all be freelancers in the future. The 27-year job like I held seems to be disappearing with the daily dead tree newspaper.

On my first job in 1970, we used manual typewriters. Soon, we switched to IBM electrics. We typed a code on the stories and old-fashioned pressmen–the guys in the paper hats–fed the stories into scanners. We became the first computer generation.

But I’ll never forget one of the columnists, Gil, couldn’t adjust to the electric typewriter. He became frustrated. He gave up. He was like the film stars of the silent era who couldn’t make the transition to the talkies. Gil retired in his ’50s.

Gil is an object lesson for us all. Unless you just want to go fishing full-time–maybe for good for you, but not me: Be open to change. Embrace new technology. Be excited about what you’re doing. Keep learning.

LTB:  But where is journalism going these days? I see that classical media is shaking and journals are changing habits. So journalism is changing too. But my specific question here is, how will scientific publishing change? We see the waves of Open Access Journals Movement (PLOS and the others) but these are still good classical peer reviewed journals. Do you see any novel type of media that could replace journals? What is your opinion about JOVE (Jounal of Visualized Experiments). Could this be a new type of publishing?  Could someone build a scientific blog and stop counting the Impact Factors but start to count visits and links? Could this kind of change happen?
Too many questions again.
Maybe let’s summarize it in one single question:
Where is scientific journalism going?

HW: Good question. But I am no visionary on scientific publications, let alone lay publications.

Maybe 10 years ago, I saw a collision coming. I saw how all lay media were becoming one. I heard audio clips on the Wall Street Journal site, and saw text stories on TV websites and now video clips on print news sites.

I even have become a bit of a video reporter. I always try to do video interviews via webcam for my stories on the Skype blog. People have different ways of accessing/inputting information. The more choices you can give them, the better your odds of reaching them.

I also took a visual story telling class to learn what I could about doing video for the Web.

That said, I am not familiar with the video project you mentioned. I’ll have to take a look.

All media seem to be going through a revolution, seeking new models and new ways to find and pay their way.

The students in my science/health writing class at Medill/Northwestern—the next generation of media— are required to do text stories. But also to do video and interactive graphics. They need to be flexible and masters or at least knowledgeable about all media.

I hope text–at least web-based articles, if not dead trees–will survive. Video story telling can be compelling. But I’d be skeptical that really complex stories can be told that way. Maybe I sound like a sentimental traditionalist, but I hope the word survives in print or on the web, for lay as well as scientific publications.

Thank you very much for your time and effort to share your ideas with us!

———————–

You can have a deeper insight in Howard’s work on the following sites: herehere, here and here

PCR or the Polymerase Chain Reaction

In animation, DNA, molecular biology, PCR, plasmid, Polymerase, Polymerase Chain Reaction, Replication, Taq on April 12, 2009 at 6:25 pm

When I first heard about the Polymerase Chain Reaction my first association was with the atomic bomb chain reaction. You know probably from your studies: the labile Uranium if receives a neutron it transformed to a stable Uranium isotope while several new neutrons are released. If these newly release neutrons meet novel labile Uranium atoms the reaction is amplified, more and more neutrons will be released until the system if is lost from control explodes in the form of an atomic bomb. If the reaction is under control we can produce heat and through this electricity in an electric plant, if the critical mass of the labile isotope is ignited with a neutron beam, it will explode.

You can have two excellent representations of the chain reaction below:

The chain reaction

But this is a blog about techniques in the molecular biology lab, so we will not deal with the fission chain reaction but we will see how a similar type of amplified reaction is produced with the DNA by specific enzymes in a reaction tube. The enzymes that can do a chain reaction are the Polymerases.

What is the function of polymerases? We have them in each of our cells. They do the most basic reaction that keeps life going on from the start of the very first organism ever. They are duplicating in a semi-conservative way the DNA in order to allow the transmission of the genetic information during cell division.

You can have an animation about DNA replication below.

What is the Polymerase doing?

The two DNA strands are connected by hydrogen bonds and code for the same information by the A:T and C:G base pairing.  The two strands are anti-parallel if we look to the double strand from one direction one of the strands will be in 5′-3′ direction and the other vice-versa in 3′-5′ direction. As an important rule we have to know that in nature all polymerases are doing the DNA synthesis in the 5′-3′ direction. The strand that can be replicated according to this rule is the leading strand. The other one is called lagging strand. The problem is that both strands have to replicated in by the same protein complex! But how can one single Replication complex produce the leading strand and the lagging strand in the same time ? We arrived to the problem of the the Okazaki fragments.

In the animation below you can find a good representation about how a single Replication complex can do the synthesis of the two different strands.

In order to speak about PCR we have to go out to trip to check some Geysers.

So let us check first the big Steamboat Geyser.

Yellowstone Steamboat Geyser

If we go closer to one of the hot springs we might see that the water is “living”, there are some algae, micro-organisms in this water. Let’s have a look:

The Yellowstone Hot Springs

These micro-organisms are living in really hot water. But if they are living, they should replicate, and if they replicate, they should have DNA polymerases!

These micro-organisms were isolated and one of them, called Thermo aquaticus (sometimes named Thermophilus aquaticus) became really famous. It has a polymerase that is used in vast majority of the in vitro DNA replication processes and in PCR.

I am sure you all have an idea already about PCR. We put all reagents needed for a DNA replication in a tube and reproduce the normal DNA replication process. So what do we need? We need a DNA template an oligonucleotide as a primer, the building blocks of the DNA (dATP, dTTP, dCTP, dGTP or in general dNTP -deoxi nucleotide tri phosphate), Mg and the Polymerase. If possible from Thermo aquaticus, which is called Taq.

There is one trick! This one trick was invented by Kary Mullis and he received Noble Prize for this single idea. The trick is, that we will not reproduce completely the natural reaction. We do not want to bother with leading strand and lagging strand and with all kind of Okazaki fragments and helicases and ligases.

The idea of Kary Mllis was that if you separate the double strand and design two oligos that will bind the two different strands but will look towards each other, than the product will be doubled. If you separate the strands by heating the solution to 95C you can repeat the reaction, and now you will have 4 copies. In the next run 8 copies and so on in each reaction you will have 2 on the power of the “cycle number” copies in a chain reaction fashion!!!

Let us have a really simple and good introduction to the whole procedure in the next two animations:

Below you can find a more fancy animation of the same procedure:

For this idea Mullis got the Noble Prize. His work changed completely the history of molecular biology. Let us check an interview with him about how he discovered PCR:

What is the practical use of this whole method?

Amplifying DNA by PCR became one of the most widely used method in a molecular biology lab. You can use it for transferring DNA from one plasmid to a different one, to introduce mutations and even to measure the amount of a specific gene in a sample. By combining it with a Reverse Transcription reaction we can measure copy numbers of RNA molecules.

Below we can see an example of how it is used in criminal justice!

In the next video you can see the workflow of DNA sequencing with a New Generation sequencing machine. What is remarkable here is that the designers of the instrument are skipping the cloning of the DNA fragments into plasmids and amplification of the plasmids by bacteria. They use micro reactors in the form of an emulsion PCR. One oligo is on a bead and the DNA binds the oligo. Each bead is fused with a small droplet that contains all the other reagents for the PCR. By this trick you will have a clonal amplification of the DNA fragments. One bead will contain on  type of DNA and you skipped all bacterial work. The result is that you can sequence the whole human genome in a couple of weeks for less the 100k USD. Or you can sequence a bacteria in a day…

Emulsion PCR in the FLX sequencer workflow

At the end of this lesson, lets have some fun and see the celebration of the PCR!

Restriction Enzyme Resources

In DNA, plasmid, Resources, Restriction Enzyme, Uncategorized on April 6, 2009 at 8:46 pm

Dear Colleagues,

Sometimes it is good to have links that cover a topic. This is why I have collected here a bunch of links that might be useful for you in your work.

You can have a good description of the methods used in restriction enzyme analysis here: Methodbook.net

If you would like to use restriction enzymes for your work, you can find a list of links of the best known restriction enzyme providers below.

New England Biolabs

Promega

Roche Applied Science: Benchmate

Invitrogen

Fermentas

If you want to start your work with these enzymes, please consult the protocol provided with the enzyme or check it at the website of the manufacturer.

Be sure you know what an isoschizomer is, what star activity is, or how you can make double digestion (details here and here).

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