Like a kid on Christmas Eve

First off I just want to clarify the title. If anyone was wondering, Christmas Eve is the major event in Swedish Christmas celebrations, and not Christmas Day. That was today's bit of cultural knowledge for you. So now on to what I actually intended to say with this blog post.

A bit more than a week ago I was doing some major lab work, which unfortunately didn't work very well. In the end I narrowed down the issue to an expired kit of reagents.
I proceeded to talk with my supervisor about this (who is in possession of the big bag of money), and she supported my decision to buy a new kit.

However, just ordering the damn stuff was not as simple as your everyday internet shopping for clothes, the latest blockbuster on blu-ray or any other odd item from a dark corner of the internet (because I'm sure even that would have been easier).
After finally finding the product I wanted, in the less than straightforward online store - I mean where have you even bought anything online where a simple search function in the store has given you nothing, but a bloody google search did? - I, as a first time customer, had to register for an account.

I've been using other online services of the same company before and was convinced that I already had an account. Oh boy was I wrong. Sure enough, I found my login credentials, but apparently the web store had a separate login system. However, instead of telling me something along the lines of  'wrong password and/or username' or 'there is no such e-mail address in our system' when I tried to retrieve the password I thought I had changed and forgotten about, their webpage simply said that the webpage I'm trying to access could not be retrieved.

Stupid me didn't realise that making another registration was even an option until two days later, when the mysterious internet access failure was still there.
After registering and adding "new" delivery and billing addresses everything had to pass through manual confirmation by the company before they would send me anything, naturally.

I thought the day I would be back in the lab would never come, in my wait I even had time to realise that I need a plan B for my current method in case everything goes south, and proceeded to order another kit (for a different but related purpose), which for the record (and thankfully), was much easier and faster to order.

So yesterday three packages with my name on them finally arrived and with all these cool new kits and reagents I felt like a kid on Christmas Eve! They were all neatly packed with two styrofoam boxes and one cardboard (there was a styrofoam box inside the middle cardboard box). The styrofoam boxes had the reagents which had to be shipped cooled or frozen, so naturally they had ice packs and dry ice in addition to my stuff.

For those that don't know anything about dry ice, it's basically frozen carbon dioxide (yes, the gas!) which has an approximate temperature of -80 degrees C.
Apart from being a convenient cooling agent it can also be used for some silly things in the lab with a little help from sample tubes, a regular sink and a little imagination.
Put a pellet of dry ice in a 2 mL tube, shake it briefly and throw it to the ground and you have your very own "fire cracker" (minus the fire). The thawing pellet will evaporate into carbon dioxide gas, build up the pressure and then pop the lid on the tube.
However, chances are that you will grow bored of cleaning up your messy floors long before you run out of dry ice pellets, so before putting away the box you could pour some of it into the sink and pour some regular tap water on it. This will quickly turn your sink into a "bubbling witch's cauldron".
There certainly are potential for some good pranks here, but if anyone asks, I never said (or did) any of these things.

Due to popular demand

In general, most of my lab oriented work has come to a grinding halt. Which is more than a little frustrating considering that my microarray slides are waiting in storage at the KI analysis facility (BEA). I'v narrowed down the issue to expired reagents as the source of my problems in quality controlling my RNA. For that reason I will not extract any more RNA, or do any other RNA work until I've received and tested the new RNA analysis kit I just ordered. Anything else would be a waste of my precious time and samples. Apart from that, I've gotten some kind assistance from my colleagues in trying out different RNA extraction kits and methods to see if I can optimize my protocol for a higher yield of good quality RNA.

On a more positive note, me and my good friend and colleague Tom got invited the the departmental plant ecology meeting to hold a condensed seminar on the first session of the workshop we hosted in communication, feedback and presentation techniques. The emphasis will be on promoting a better 'feedback-culture', since most of us agree that the scientific community could do good with a lecture on how to communicate properly.

I feel honored to have been given this opportunity and I'm hoping for a large, enthusiastic and diverse audience so we can spark some interesting discussions.
I also feel that this is a testimony to the success of the workshop, and depending on the outcome of the seminar we could be looking at another workshop next year. Likely the same format, but with some fine tuning and improvements based on previous participant evaluations. However, it will only happen if we're asked (read begged), since both me and Tom are in no need of extra credits (HP) and are only doing it because we like the subject.
Stay tuned.

How was it, being on national TV?

In the end I got a lot of praise for my attendance in the SVT Barnkanalen Morgonshowen kids show. Everything from a simple 'well done you!' to a friendly suggestion of a career change (to TV host).
However, funny thing is that it was more or less chance that gave me this opportunity. Because that's exactly what it was, despite "only" being a kids show. An opportunity to communicate science.

Basically an editor at Morgonshowen called a professor at my department in search of microscopes and someone who could show them, and small things in them, in their show. I've no idea why they didn't contact the the Biomolecular department, but that doesn't matter.
The professor they first contacted didn't have access to any microscopes and those research groups who did included mine and a few others. Mine just happened to be the first one to be contacted and since I'm the only swedish speaking researcher in my group that meant that I would be the only one capable of pulling it off.

Before I even knew what the fuss was all about I was reading through a brief manuscript on looking at small, everyday things in a microscope. Apart from that I was free to add my own research and expertise. Knowing that, the decision was a no-brainer and in hindsight I think it turned out even better than I had hoped.
Being a perfectionist I immediately started to put together an instrument/computer setup that would allow me to best visualize what I wanted to show, and of course, writing a simple manuscript of my own with simple facts and metaphors that would come in handy during the actual recording in their studio. During that time I also, in my mind, went through what I would say in response to different questions I would get, since it would mostly be improvisation at the time of recording.

In the end I brought two different microscopes (40x and 400x magnification), a camera that fit both microscopes, my laptop with visualization softwares and a 24 inch monitor. The microscope slides and materials were all prepared in advance and with the helpful assistance and support of my supervisor I had everything ready, including tiny beakers with phytoplankton cultures of different colors (e.g. red, green and pink), within two days. Despite these preparations there were still some improvisation happening with the microscopes on the day of recording.

The setup I used, trying it out in my lab (one microscope less).

When that day finally arrived I was really nervous. I remember that my supervisor and the public relations manager at the department accompanied me to the studio, and the drive there seemed to last forever. I had the feeling that I wanted to get to the studio that very instant and just get this thing started so I could better handle my nervousness.
The security at the SVT main building was rigid and we had to register and wait for quite some time before the editor finally showed up to greet us and let us into the studio.
My setup was already there and ready to go, since I assembled everything the night before. All I needed to do was plug in my computer. But before that happened, I met all the people involved, including of course, the host of the show for this episode, Tobbe 'Trollkarl'. Those of you who aren't swedish will probably not know who he is, but suffice to say that he has been on TV for a relatively long time, even when I was a kid. So I had a surreal moment of meet and greet with him before getting ready. The studio was fairly large and had a double setup depending on the different shows they were recording. It was a ridiculous amount of stuff, both technical and otherwise, lying around waiting to be put to use. And of course, in the middle of it all, a massive mixer table, cameras, lights microphones, you name it.

Apart from double checking my instruments there was not much preparation to be done. No make-up, no briefing, I just put my lab coat on and had a brief rundown with Tobbe about what he was going to say and do. I more or less played along and did my thing. To be honest it was a fairly simple task since Tobbe made life easy for me with his genuine interest and excitement over getting the opportunity to look at all these different things in a microscope. For example, I would guess that a lot of people have no idea that grains of regular salt is actually cube shaped.

Me and Tobbe Trollkarl in the studio

The recording was done in three segments and during the first I was really nervous, and I remember my supervisor happily saying that I should try and smile more. In time for the second segment I was getting warmed up and everything felt more comfortable. I got some pointers from the camera woman which also helped. In the end it felt really good, even though I always get the nagging feeling that there are things I could have done better, which I guess is natural.
By the time we did the third segment I didn't want it to end. I could do this for quite some time more. I had a blast.

There were no re-recordings and the session, which should have taken two hours, was done in only one hour. Everyone seemed really pleased with how it turned out and after watching the show on TV on the 27th of August I can see why, although it's quite weird seeing myself on TV. My kids were thrilled!
The week after the recording in the SVT Barnkanalen studio the editor called me again.

RNA extraction and quality control of your Volvo

I got an e-mail the other day. It was from Agilent who said that my order of two custom microarray slides scheduled to arrive around the 5th of September were already finished and were being shipped from Germany to the analysis facility BEA at Karolinska Institutet (KI) now.

Guys! You're two weeks early! I haven't even gotten my own stuff (samples) together yet. Germans... How often does this happen? By experience - always when you're lagging behind. That's not what I replied though, but it's what I silently told myself.

Now you might think 'hey, you only have to fill one slide, 8 samples, how hard can it be?'. If you just thought exactly that, I'll give it to you. You're right, it's only 8 samples and it would take me 2 hours to extract. However (I hate that word, and I use it all the time), out of the 12 samples that were already extracted only 2 had enough total RNA for analysis, and don't get me started on the quality control of that RNA... ops, I already did that myself... they all failed.

At this point I have nothing. Fortunately for me, we have lots of archived RNA with corresponding DNA samples which have already been processed in the qPCR. Basically, that means that I can see in which RNA sample that I should have the right organisms and enough material. However (here we go again), that doesn't guarantee good enough quality RNA.

There are several reasons why the RNA in my samples might not be of good enough quality, and when I'm talking about quality it's rather specific and doesn't entail the comparisons between e.g. longevity of brands that we're so used to in modern consumption society. No, here, I'm talking about degradation and size of the RNA fragments. Like if you removed a few parts from your Volvo, it would still be a Volvo, but it would probably not function very well, and if the parts removed were body parts, the untrained eye might not even recognise the car as a Volvo anymore.
A similar problem arise when I have low quality (degraded) RNA. The probes in my microarray are all extremely specific (untrained eyes) and when the genes (Volvos) in my samples are missing a few parts due to degradation they will not be recognised.

Image credit: Fabian Oefner

To be short and simple, RNA is the coded message that our DNA relays for our cells to, down the line, translate and produce proteins with various tasks and properties. Naturally, the message doesn't stay intact for very long since, in most cases, when the message has been delivered and translated it has outlived its usefulness and is recycled within the cell.
So, I might just have been unlucky and the RNA in my samples was naturally degraded when the research vessel arrived (too late) on site. I might do a terrible job extracting the RNA, and perhaps, mysteriously introduce RNase (an RNA degrading enzyme) to my samples. The agitation step (shaking the samples really, I mean REALLY, fast together with tiny glass beads to crack the cells open) might shear the RNA to tiny bits and pieces. The latter is the only explanation that I can actually test.

I'm assessing the RNA quality in an instrument called Bioanalyzer 2100 (Agilent), which gives me a value from 1 to 10 (10 being the best - most intact RNA). It has developed into an industry standard since it uses an algorithm to objectively calculate and compare the amount of two different ribosomal RNAs. This way us scientists know that we will get good and reliable results and we secure the possibility of reproducing the experiment (important for e.g. peer-review and validation). Everyone is happy. Me included, if I can get good quality RNA, otherwise no.

Grit and the growth mindset: The recipe for success

I recently watched a TED talk by Angela Lee Duckworth. It was recorded in 2013, so I have no idea how this inspiring talk has eluded me for so long.
Anyway, Lee Duckworth tells her tale of how she left a demanding but successful job in consulting to teach math to 7 graders. That's where her ideas and research started to develop when she observed the struggles of her pupils. After a few years she started studying psychology and pursue the unverified observations she made as a math teacher.

You can see the entire 6 min talk right here.

In her talk she also mentions, at the very end, a concept called the 'growth mindset'. This is also an idea that has been around for quite a few years. It was coined almost 30 years ago by Carol Dweck who studied how students dealt with failure. The opposite of the 'growth mindset' is the 'fixed mindset' and what sets them apart is the belief that talent is either static or developmental. This means that if you believe in the 'growth mindset', talents, abilities and even intelligence is not something that is restricted to life's lottery, but is actually something you can improve and develop.

I first heard about the 'growth mindset' during the workshop I co-hosted in Communication, feedback and presentation techniques, where I had a very interesting discussion with my department's Public relations manager. I didn't think much more of it after that, to be honest, but when I now heard it in the context of Lee Duckworth's TED talk, it made perfect sense. It is also how I would define my own success.

So together with the 'take home word of the talk': grit - the 'growth mindset' is what defines success. As nicely put by Lee Duckworth "grit is living life like it's a marathon, not a sprint". It's about setting up long term goals and sticking to them, not by hours, days or weeks, but years. Perseverance.
It's about not giving up, and use the inevitable failures to grow, work harder/smarter, and eventually succeed. Because the thing about success is that it doesn't always happen right when we want it to happen.

Lee Duckworth wraps up her talk with the task that stands before us all, and that is teaching our kids to be grittier. Because talent and intelligence means nothing if you're not willing to work hard.
When I was a kid I played hockey and I strongly remember a small piece of paper that hung on the whiteboard in the club's meeting room. It said: "The will to win must be stronger than the fear of losing". I didn't quite understand the deeper meaning of it back then, and I'm not sure whoever put it up did either.

In hindsight I wish that someone would have explained it to me. I wish that I had even a fraction of the knowledge I have now, back then, and I'm surely not alone with that wish.
I think Lee Duckworth is right, and when I think more about it, I'm actually subtly teaching my kids grit. Perhaps it's time that we do the same, although more formally, in our schools.
No matter how small the task I always tell my kids to never give up and keep trying until they make it. Often to their utter frustration, when they can't get their seatbelts fastened in the car. However, not a single occasion have they not made it, eventually, and their howling annoyance been instantly replaced by silence. Dad was right.

Cross-hybridization

This is going to be a tough one, and I'm afraid I might end up with more questions than answers at the end of this blogpost. Either way I feel like I need to clear my mind on the matter of cross-hybridization.

After a long and nice summer holiday with my family, work didn't progress much, even though I had hoped that my external contacts would be able to do something after final submission of my microarray design. However, it turned out that everyone else was gone on their summer holidays too. I should have known.
Now everything is moving along nicely again, but the road up to the final design was certainly not a straight one, mainly due to cross-hybridization.

This issue has been a constant grain of sand in my eye since the day I set foot in a molecular lab, however, it got all the more profound when I started working with qPCR and gene expressions.
As of now, there's no satisfactory way of dealing with it, we just know that it's there and can do our best to account for it in our data analyses and interpretations.
My next project (after the microarray) will attempt to at least improve (decrease cross-hybridization) for some of the known and widely used oligonucleotides in marine cyanobacterial diazotroph research.

Cross-hybridization is basically hybridization of a primer or probe sequence with a sample sequence other than the intended target. Often it happens due to several sequences being highly similar, but also because the oligonucleotides used are not specific enough in their design.
What I mean with specificity in this case is that even though your oligonucleotide design is a perfect match to a certain region of a gene in a target organism, it could match almost, if not just as well, for a closely related species with the same gene. Moreover, even a close match (not perfect) could be enough for cross-hybridization to some degree. There are several reasons why mismatches in nucleotides still hybridize, like for example the nucleotide content of the target sequence (G-C-T-A) and the location of the mismatch in relation to the 5' and 3' ends.
To make matters even more complicated, the concept of species is kind of blurred when working with prokaryotes, where we often are interested in different strains of the same species. Even though they're systematically the same species, phylogenetically they're not, and potentially have different impacts on different ecosystems.

So running cross-hybridization tests where I use oligonucleotides designed for a particular target organism on a closely related strain or species to see if I get a signal. If I do get a signal, it is a cause for concern, but if I don't get a signal it is safe to say that the designed oligonucleotides are able to distinguish the target organism from at least that particular closely related strain or species which I tested against.
Of course it is impossible to test for every single possible cross-hybridization. There are simply too many unknowns involved and for some strains we don't even have any designed oligonucleotides yet. It is also possible that there are closely related strains or species out there in the ocean that we just don't even know exist.

An emerging number of marine cyanobacterial diazotrophs are just known by sequence, matched by phylogeny, in metatranscriptomic studies. My diatom diazotroph associations (DDAs) for example, are easily observable in a fluorescence microscope, so I can quickly look and see if they're there and which species and strain it is. That will initially be key to improving and optimizing oligonucleotide designs for the larger marine cyanobacterial diazotrophs. For the smaller we're somewhat dealing with a black box and I'm expecting a trial and error approach.
Up to this point we've mainly focused on the nifH gene for our target sequences, but as these organisms get fully sequenced, we might find alternative genes which are better suited for our purposes, based on relatively subtle genetic differences.

Seminars and presentation techniques

I thought about writing a blogpost about cross-hybridization after my last post which barely touched upon the tricky issue. However, two rather technical blog posts in a row might be a bit too much to digest.

So, instead, after having two scientific seminars (about my research) in three weeks, I decided that it would probably be a good idea to write about my experience and thoughts on my presentations and also presentations in general.

I feel that I've talked enough about my licentiate seminar, so I will kick off by mentioning the nice introduction I got from my PhD colleague at the beginning of my last seminar. I was fairly well informed about the content of the introduction (since he asked me about it), but one thing that still got me thinking was when he mentioned that I co-hosted a PhD workshop/course on communication, feedback and presentation techniques earlier this year. It was very successful (based on participant evaluations) which opened up for further development and a chance to host it again next year.
One of the most common questions that I got during the workshop concerned a list of recommendations, or do's and don't's, of oral presentations. I was a bit reluctant to answer this question, especially since we touched relatively little on this subject in our theory and the presentation techniques were mostly based on self-development and feedback from the participants. I also believe that there is not one single recipe that fits everyone in this case, rather I think each and every speaker have to find their own personal style and the tricks that make them comfortable standing in front of a crowd.

So, where am I getting with this? Well, after my two recent seminars, and getting lots of positive feedback from both fellow PhD students and faculty, I might not be giving myself enough credit in terms of my knowledge in presentation techniques (and perhaps even my research).
I think anyone who has ever acquired a Masters degree (or similar achievements) is familiar with the question: "what the have I really learned during these years" when looking to, for example, apply for your first job.
Sure, I have no formal training in oral presentations, although I do have a neat little hardcover book in my office on how to improve your oral presentation skills, but I haven't even nearly finished reading it. But practice makes perfect, right? And with enough experience you're sure to at least know a few tricks of your own. However, never underestimate the importance of watching and learning from other great speakers. They are great because they had the courage to walk that uncomfortable path of trial, error, feedback and improvement.

With that said I would like to share my experience, feelings and thoughts on the recent seminar and oral presentations in general. Perhaps some of it might be incorporated into the next workshop. Who knows.


1. Preparations

A good presentation is based on good preparations. Of course this entails having a solid story and/or material to talk about. In my case, an overwhelming majority of my presentations use visual aid like a projector and PowerPoint. Visual aid, if used right, is a powerful tool, and it can also be used to direct a staring (and perhaps sometimes intimidating) audience away from you. However, is that always a good thing?
When you have your presentation set out and you know your format, we come to the more practical preparations. Personally, I like to set up, practice and get a feel for the lecture hall of my presentation beforehand. This way I can make sure that my slides work, I know if there are any pointers available and I can get a 'general feel' for the space that is my stage. However, when it comes to e.g. conferences, this might not be possible, so bring your own gear just in case.

2. Practice

Without visual aid, practice becomes even more important since you can't rely on any slides to help you back on track if you wander off into a dead end. Also, if English isn't your native language it could be even harder than usual to improvise even though you know your story very well.
So practice is key, but it is important to separate practice from learning. Learning your talk is what you do in front of your computer, manuscript in hand perhaps, while practice is what you do 'on stage'. However, when you practice you chose both the setting and the stage. It could be talking to yourself in the mirror or gathering a group of friends as audience in an otherwise empty lecture hall. In preparation of my licentiate seminar i practiced in the actual lecture hall of my licentiate seminar, with and without an audience, every day for two weeks.

3. Feedback

Unfortunately feedback is often overlooked or confused with criticism, and both givers and receivers are responsible for this somewhat lingering mentality.
So from now on, stay away from the word 'criticism' and instead use constructive feedback, both when giving and receiving. Constructive feedback can in turn be either positive or negative, but it's still constructive! Constructive as in a tool for improvement. Because it is important to highlight both that which can be improved and also reinforced.
Feedback could be uncomfortable at first, so bring your friends and close colleagues to your practice talks.
I specifically asked for feedback at three different practice occasions from three different audiences (background, expertise, age, gender, etc.). All their different feedback significantly helped me hone in on the kind of talk and message I wanted to deliver to my intended audience.

4. Consistency

Being a consistently great speaker is unfortunately not just finding your comfort zone and blast through everything you want to say to every audience you will ever have the fortune to stand in front of.
Even a comfortable speaker will have to reflect upon the message being delivered and how to deliver that message. Most, if not all, depend on the audience and the setting. Are you presenting for your peers or a group of school kids? Even if it's an audience of scientists, do they know your field as well as you or your grandmother? Are you the first speaker or the last speaker of the day? Both scenarios might mean that you will have to try extra hard for their undivided attention.
Simplifying and explaining important concepts in detail helps, but don't be too long, and when people start to look tired, change your pace, voice tone and/or body language.

5. Body language

Right here is where your talk will be decided as great or not. Body language. It will not only affect your stance on stage, but it will also affect your voice, your breathing and your connection with the audience.
When I attended the introductory course in research studies in biology during my first year as a PhD student, we learned about something called the 'power spot'. It is not only a certain piece of ground, but also a stance by you, where you are at your most comfortable, confident and connected with your audience. Basically, you stand up front, on your two firm feet, looking the audience in the eye. If you can find your 'power spot', you've already come a long way. The opposite would be standing hunched behind a podium, looking at your own PowerPoint images the entire talk. We've all seen it, and know that it makes people cringe and lose focus.
In your 'power spot' you own the stage and the moment and even though you might be battling your nerves at that very moment, you will still radiate confidence. Time your talk with a few well placed breaks and allow yourself to breathe and the battle will be won. In my last talk I had shorter breaks (just breathe) of a few seconds and then three longer planned breaks where I took the time to zip some water. Trust me, it will help. I asked for specific feedback on this behaviour and everyone I asked thought of it as a sign of confidence or they didn't even notice. Sometime even the audience could use a breather.

6. Patience

After reading this blogpost, even after reading a whole book on the subject or watching a hundred TED talks, there will still be things that could be improved. This list is by no means comprehensive. If you want to be serious about nailing that perfect talk, set up goals along the way. Watch other great speakers and practice different approaches. No speaker woke up one morning, stood up in front of an audience and delivered the most mind-blowing talk ever! It will take time and effort, and likely it's a journey without a final destination.
Personally, I will likely never be fully rid of my nervousness prior to a talk, no matter the context or the audience, but I've learned how to deal with it, and if I compare to where I started I've undoubtedly come a long way. I was on the verge of throwing up prior to talks in high school - where did you start, and where are you now?