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#RealTimeChem Week 2015 Awards and Cook Off prize winners

Hi RealTimeChemists,

Once again I have to say a massive thank you every single chemist out there who took part in #RealTimeChem week. This community only exists because of you and it’s always great fun to witness the massive variety of chemists taking part from all over the world. It was nice to see tweet not just from regulars, but also a lot of newcomers. Welcome to #RealTimeChem I hope you enjoy your stay.

While the week event is over for another year, don’t forget that #RealTimeChem is a 24/7 project, so feel free to keep sharing chemistry whenever you want and engage with your fellow chemists around the world.

I have some very special thank yous to give out this year to the following folks:

  • Andy Brunning of @compoundchem fame. Not only did Andy design all of the graphics for this year, he also provided a really cool infographic competition of his own to celebrate #RealTimeChem Week. I look forward to working with Andy again in the future.
  • Katey Birtcher and all the folks at Elsevier’s SciTechConnect who ran the Blog Carnival this year and promoted the week. Your enthusiasm for the project has really kept me going and I loved the round ups.
  • Nicola Burton, formerly @Elements_UD now @SpaceBambee, thanks greatly for the #RealTimeChem Award badges
  • Kudos to the Royal Society of Chemistry and all my friends at work for supporting the project and also getting on board from time to time.
  • Finally, thanks to Guido Kemeling, Editor-in-Chief of @ChemSusChem, who was kind enough to organise the prizes for the The Great #RealTimeChem Cook Off. Thanks to all the other ladies and gentlemen at WileyVCH for the support as well.

With all of these out of the way, it’s time to get onto this years awards. This was, as ever, ridiculously tough because there were many excellent tweets all deserving a prize. Thankfully, I have a few more prizes to give out this time around!

Below you will find out the results. Drum roll please?




The three tweeters below all win a #RealTimeChem Week 2015 mug of their very own like the below:

Everyone loves a mug right? You can do all kinds of things with them!


From Wednesday. Brian Wagner (@drummerboy2112) is one of THE chemists to follow on Twitter in my humble opinion and this simple demonstration of Boyle’s Law using a marshmallow, a flask and some suction was arguably the most popular tweet of the whole week in terms of retweets and favourites. Science can be fun and informative both at the same time.


From Wednesday. Laura van Laeren (@lauravlaeren) strikes again, sharing a lot of great tweets throughout the week, including several pictures of her beautifully painted finger nails. However, it was her starry night flask that caught a lot of attention this year and I couldn’t ignore it’s awesomeness. Pretty colours for the win!


From Saturday. Speaking of pretty colours, it was National Chemistry Week in the US at the same time, with colours being the big theme. Emily Hardy (@EmilyEHardy) snuck in on the Saturday to show off some fantastic chemiluminescence. Really spectacular.

Don’t forget to DM me your address so I can post your prize to you.


The remaining 9 Pt Awards winners will all get a #RealTimeChem keyring like the one below:


All winners, DM me your address & I’ll post your key ring to you.

NOTE: Au/Ag award winners. Unfortunately, you don’t get a prize, but I shall in the near future send you a small certificate of achievement (via Twitter) for you to keep, one for each award you received. 


I was lucky this year to be given some extra prizes for a new competition. The first #RealTimeChem Cook off! There were quite a few entries and below you can find the 6 chosen winners.

The Grumpy Chemist (@Chemistry_Kat) 

Henrik Pedersen (@hacp81)

Victoria Stafford (@ToriaStafford)

Massimo Grillo (@MassimoGrillo63)

Tom Kuntzleman (@pchemstud)

Debbie Mitchell (@heydebigale)


All 6 of these tweeters win a copy of “What’s Cooking in Chemistry?: How Leading Chemists Succeed in the Kitchen“.

Send me your address details via DM and the folks at Wiley will send you your prize*!


*Book prizes sponsored by @WileyVCH’s society chemistry journals. These are: @ChemEurJ., @ChemistryOpen, @ChemBioChem, @ChemCatChem, @ChemMedChem@ChemElectroChem, @ChemPhysChem, @ChemPlusChem, @ChemSusChem, @EurJIC, and Eur. J. Org. Chem (all journals of ChemPubSoc Europe); @ChemAsianJ, @AsianJOrgChem, and @ChemNanoMat (all journals of the Asian Chemical Editorial Society); and @angew_chem (a journal of @GDCh_aktuell)



Well, that’s all folks. All awards are now given and #RealTimeChem Week is officially over for this year. Once more, thank you to everyone who took part. May all your chemistry dreams come true.

mischief managed

-Doctor Galactic & The Lab Coat Cowboy-

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“The Great #RealTimeChem Cook Off” Competition


Chemistry is like cooking. Just don’t lick the spoon.

Hello RealTimeChemists,

Over the past few years I’ve noticed that many of you have shared some fantastic creations, not just from your laboratory at work, but also from your kitchen at home.

There is a very strong association between chemistry and cooking, and to celebrate this connection, I’m very happy to announce the first ever #RealTimeChem Cook off!


Don’t worry these guys aren’t going to be judging you. © BBC

Prizes for the Cook off have been kindly provided by the folks at WileyVCH* [special thanks to Guido Kemeling (E-in-C of @ChemSusChem), @ChemSusChem@angew_chem & @AsianJOrgChem). We have 6 copies of “What’s Cooking in Chemistry?: How Leading Chemists Succeed in the Kitchen” to give away. This fascinating book acts as a Who’s Who in organic chemistry, showing what top scientists like to cook.

All you have to do to enter the competition is to tweet a culinary creation (anything cooking, baking or food related) using the hashtags #RealTimeChem & #whatscooking this week. Your tweet should ideally include a short description with a picture or video of your creation (the description can even talk about the chemistry in your cooking! It’s up to you). Alternatively, you can write a recipe for others to try. 


Everyone who Tweets a cooking related tweet using these hashtags will be entered into the competition and 6 favourites will win a book. This competition runs until the end of #RealTimeChem Week 2015 (25th October) so you’ve got some time to get some ingredients in and post a tweet (I mean cake baking is such a weekend thing!). I expect many of you will have some free time to get in the kitchen on the weekend in particular, so the perfect excuse to share a little more fun #RealTimeChem and possibly win a prize. Just remember Mary Berry expects…

mary berry


If you have any questions about the competition, please let me know via Twitter (@RealTimeChem) 

-Doctor Galactic-

*Book prizes sponsored by @WileyVCH’s society chemistry journals. These are: @ChemEurJ., @ChemistryOpen, @ChemBioChem, @ChemCatChem, @ChemMedChem@ChemElectroChem, @ChemPhysChem, @ChemPlusChem, @ChemSusChem, @EurJIC, and Eur. J. Org. Chem (all journals of ChemPubSoc Europe); @ChemAsianJ, @AsianJOrgChem, and @ChemNanoMat (all journals of the Asian Chemical Editorial Society); and @angew_chem (a journal of @GDCh_aktuell)

About the #RealTimeChem Week 2015 Awards

Hi everybody,

#RealTimeChem Week 2015 is almost upon us and I’ve had a few folks on Twitter asking me about the Awards that are given out during the event. Information can be found on the FAQ like everything else about #RealTimeChem, but I thought I’d make things even clearer in a separate post.

What are the awards?

Every year I bestow awards upon my favourite contributions to #RealTimeChem Week. This year’s awards have been revamped a little with a new structure and some extra prizes for those winning the top awards.

Awards are given out each weekday with a joint weekend award giving you 6 opportunities to win.

There are three levels of award: silver, gold and platinum as shown in all their shiny goodness below:


The new award badges this year have been provided by the excellent Nicola Burton (formerly on Twitter as @Elements_UD) creator of Elements Unleashed.

Overall there will be 12 awards given each day: 6 silver, 4 gold and 2 platinum.

How do I enter?

Simple. Tweet some fantastic chemistry any time between 19th-25th October using #RealTimeChem in your tweet and you are in with a chance of winning.

Please note: While you can most certainly win multiple awards throughout the week, in the interest of fairness I try to make sure that Tweeters only win one Platinum Award during the week. This is in order to give more folks the chance to win a prize.

What can I win? 

The prizes for the #RealTimeChem Week awards do come out of my somewhat meagre pocket so unfortunately I can’t give prizes to every award winner , but everyone will each receive a PDF certificate to keep for posterity (Silver, Gold or Platinum).

Winners of the platinum awards each day will also have the chance to win the big prize as one of the top 3Tweets of the Week”. All 12 winners of platinum awards will be entered into the final running for “Tweets of the week” with the 3 winners receiving the top prize and the remaining 9 platinum award winners receiving the smaller secondary prize.

This year’s prizes are shown below and are adorned with this years logo designed by @compoundchem.

Tweets of the week top prize: 

Everyone loves a mug right? You can do all kinds of things with them!

Everyone loves a mug right? You can do all kinds of things with them!

Platinum awards secondary prize:

A snazzy keyring for the remaining 9 Pt award winners (in case you are wondering it’s blank on the back.)

If you have any other questions about the awards, please ask me on Twitter.

-Doctor Galactic-

Announcing Compound Interest’s #RealTimeChem Week 2015 competition!

Hi everybody,

I have a rather exciting announcement for #RealTimeChem Week 2015, courtesy of Andy Brunning of Compound Interest: a competition for all you research chemists out there. This is your chance to have your research turned into an info-graphic by Compound Interest and featured on the website during #RealTimeChem week.

Compound Interest’s graphics have have been featured on sites all over the world, including The Guardian, Huffington Post, Forbes, Buzzfeed, IFLS, io9, NPR, Smithsonian, Vox, The Mail Online, and Business Insider.

See below of all of the details of this great competition:


Want a Compound Interest graphic made based on your research? For this year’s #RealTimeChem Week, Compound Interest is looking for chemistry researchers who want to explain the everyday applications of their area of research. To enter, all you have to do is write a piece no longer than 500 words long, detailing the focus of your work and its relevance to non-chemists, in language non-chemists can understand.

From the submitted pieces, five will be chosen to have graphics made based on them, and these graphics will then be featured on the Compound Interest site during #RealTimeChem Week. To enter, email your 500 word piece to realtimechemweek@.

-Doctor Galactic-

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Crystals are a girl chemist’s best friend

My name is Anna Ahveninen. Although that surname can try to convince you otherwise, I’m half a year into my PhD at the University of Melbourne, in Australia. The broad scope of my project is the synthesis of metallosupramolecules and their characterization by X-ray crystallography. The finer details? Well, that’s taking a while to figure out.


I’ve only been at the University of Melbourne for as long as I have been working on my PhD. I moved to the Abrahams-Robson group from Monash University, where I completed my undergraduate degree with honours. Having fallen in love with transition metal chemistry — the beautiful coloured complexes and their satisfyingly sparkly crystals — and crystallography in my honours year, the transition to my current project was not a difficult one. Kickstarting it has definitely been troublesome, however. In the past six months, I have been chasing a discrete assembly without a grain of success. The last two months saw a change in my focus from discrete assemblies to coordination polymers (with the same coordination motif), and just a few short weeks ago, I finally hit the jackpot. A red, sparkling, reproducible jackpot.

Since then, I have been working away at trying to turn that result into more results, hoping that it will propagate into a project and grow, with care and love and hard work, into a thesis. The following is a sample of how I am going about that.


Mondays are pretty exciting for someone working on a crystallography project. Mondays mean that my reactions will all have had at least two extra days to crystallise! I pick up my rack of vials and carry it with a flourish over to the microscope to check for clean edges and tell-tale sparkling. Since we do not have a microscope with a camera in-built, macroscopic pictures of my sparklers will have to satisfy you (Fig. 1).

Figure 1: Vials full of sparkly crystals, ripe for the X-ray diffractometer.

Figure 1: Vials full of sparkly crystals, ripe for the X-ray diffractometer.

I set about my run-of-the-mill inorganicky business until my group’s favourite time of the day: tea time. Although we have no formal group meetings, we meet with our supervisors every day around 4 pm for tea. It gives us the opportunity to ask questions of our supervisors and bring new results to their attention, while also being a nice break and group bonding activity. The group bonding consists of doing the quiz in the Herald Sun and a game involving Fred Basset. Fred is a little tradition that goes far back enough in the Abrahams-Robson group that its origins are unclear. In this game, one of our group members describes the comic strip (Fig. 2). Our job is then to guess what Fred says in the last frame. Weirder than weird to an outsider, this tradition absolutely grows on you, and has become akin to a religious duty in our group.

Figure 2: Fred Basset in his natural habitat. Fred's home is at gocomics.

Figure 2: Fred Basset in his natural habitat. Fred’s home is at gocomics.

My afternoon comes with the pleasant surprise of overnight time on the X-ray diffractometer. One of our postdocs does all of the diffractometer time allocation to ensure that the time is divided fairly, so it always seems to spring up on me.

The X-ray diffractometer (Fig. 3) has to be my favourite instrument. I get a serious thrill when sorting through crystals on a glass slide under the microscope, picking the one I think looks the most promising, mounting it on the diffractometer, centering it and then shining some X-rays on it. The excitement builds at the initial blank frame, and a few seconds later – boom! Diffraction (Fig. 4)! As is common in science, the usual result is very little diffraction, streaky diffraction, or no diffraction at all. It’s all worth it, though, when that first frame flashes up and the spots are well-defined and single and strong and beautiful.

Figure 3: The University of Melbourne X-ray diffractometer.

Figure 3: The University of Melbourne X-ray diffractometer.

Figure 4: A frame from one of my X-ray diffraction data collections.

Figure 4: A frame from one of my X-ray diffraction data collections.



The morning begins with a coffee with my group mates, followed by the weekly inorganic chemistry seminar. This week, it is a group member’s colloquium, wherein he has chosen a field of chemistry outside his project to give a talk on. These talks are very interesting to listen to and are usually very educational, both for the speaker and the audience. The rest of the day is spent trying to make sense of my X-ray diffraction data, since I have had the misfortune to be working with high-symmetry cubic systems with a high degree of disorder.

Late in the afternoon, I stop bashing my head against the crystallography wall and take some of my amorphous and microcrystalline samples to the IR spectrometer in the teaching labs. IR spectrometry is free and easy; it helps give me an idea of whether a reaction that doesn’t want to grow nice crystals is worth pursuing.


Wednesday morning is when I would usually demonstrate for my first year class, but since there are no first year practicals running this week, I get a free morning. I spend my time marking reports from the previous experiment. I turn my attention to the lab afterward, but discover that frantic preparation for powder samples for the Australian Synchrotron from two weeks prior has left my stash of 3 mL plastic syringes precariously low. I get a reaction or two in, and am then forced to find something else to do while I wait for the chemistry store to fill my order.

Mid-afternoon, I meet with my supervisor for a long talk regarding my red, sparkling, reproducible jackpot and where we can take my project from here. An hour of musing, brainstorming and me frantically scribbling down notes later, we break for tea. My spirits are elevated and the future of chemistry is looking good.


To my annoyance, I discover that the delivery of 3 mL plastic syringes is excruciatingly slow. Crippled into inability to do my reactions, I spend part of my day backing up my lab notebook. A good method that I learnt from the postdoc in my honours year, is to take pictures of your notebook pages and create an index in Excel to correspond to compound syntheses found on particular pages.

Leafing through my notebook leads to a decision to create a spreadsheet to track the variables of reactions I have been doing. I feel more secure having it available at a glance and organised, as I swear I can feel the details slipping out of my brain. I also spend some time catching up on my journal RSS feed, which I admittedly ignore in favour of doing lab work much more often than I should.


With the delivery of my plastic syringes, I can get into some serious synthesis action. My ligand, when deprotonated, tends to oxidise easily in air. To combat this, I bubble nitrogen gas through all three layers to drive out as much air as possible before layering my ligand with a layer containing a base, a metal salt and a counter-ion (Figure 5). The third vial contains a buffer layer between the two. I run two reactions parallel, as this saves me time in the long run.

Figure 5: How metallosupramolecular chemists do air-sensitive chemistry.

Figure 5: How metallosupramolecular chemists do air-sensitive chemistry.

In case you are curious, the 3 mL syringes come in during layering. I layer my reactions in the reverse order, starting with the least dense layer. Then, I inject the buffer layer below the initial solution, and finally, the densest layer. The volume of the syringes is important since I don’t like to do more than one injection per layer: for one, the suba seal becomes compromised quicker, and for another, it is easier to mess up the layering with more than one injection. Syringes with a too-high volume are also unwieldy and tend to draw in too much gas. When layered well, the reactions can look pretty spectacular (Figure 6).

Figure 6: Either layered reactions or bottled sunrise.

Figure 6: Either layered reactions or bottled sunrise.

My day, and week, draws to a close with drinks, snacks and a game of Cards Against Humanity with my group mates. What better way to end a week of brain-intensive work than a really inappropriate game with a bunch of really awesome people? It’s evenings like these that remind you that life – and science – are awesome.

Author biography

AnnaBioAnna Ahveninen was born and raised in Finland. She completed her Bachelor of Science with Honours in 2014 at Monash University, Melbourne, Australia. She is currently a PhD student under the supervision of Assoc. Prof. Brendan Abrahams at the University of Melbourne. She tweets under the handle @Lady_Beaker and blogs on Chemistry Intersection.

If you are a blogger interested in writing a guest post for #RealTimeChemInFocus, please get in touch with @RealTimeChem on Twitter.
Also don’t forget about #RealTimeChem Week 2015’s blog carnival, starting 19th October. Find out more here.
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Announcing the host of the #RealTimeChem Week 2015 Blog Carnival

Hi #RealTimeChemists,


A brief update on #RealTimeChem Week 2015 (more to come). In previous years the Blog Carnival has been hosted by @JessTheChemist on her blog “The Organic Solution”. Unfortunately, Jess has moved on to pastures new and her blog is no longer that active, so I’ve sought out a new blog to host the carnival this year.

I am pleased to announce that the folks at Elsevier’s SciTech Connect have kindly volunteered to host this the #RealTimeChem Week 2015 blog carnival. You can find a preview post on that SciTech Connect have put up on the subject here, which contains all you need to know about the blog carnival.

I am really hoping that many of you chemistry bloggers out there will join in to write a blog post for #RealTimeChem Week 2015, which starts on 19th October. Once you’ve written your post don’t forget to share it during #RealTimeChem Week with one of the two topic hashtags: #OldTimeChem and #FutureTimeChem. The nice folks at SciTech Connect will be looking out for these hashtags and collecting your blog posts together in daily round ups.


If you need a reminder of the two topics this year:OldTimeChem-PosterFutureTimeChem-Poster

Happy writing!

-Doctor Galactic-

Joining the dark side of the Force for a week

Hello! I am Clemens, a postdoc at the University of Cambridge, and in this #RealTimeChemInFocus blog post you will follow me, a chemist, doing some “biology” a.k.a. the dark side of the Force.


I know it’s weird. Why would a chemist venture into the world of biology in the first place? Honestly, it just happened! Organic chemistry was my first love as an undergrad, even after my final product of a 9-step carbohydrate synthesis decided to spontaneously decompose! I still try holding on to my first love by attempting to solve the Denksport problems of Dirk Trauner’s group and I still admire elegant total syntheses. However, the dark side of the Force has always been strong in me and over the course of my PhD and postdoc, I gradually moved towards chemical biology. I can’t help it; I am simply fascinated how chemistry can give answers to complex problems by probing or perturbing cellular systems. So, without further ado, this is a typical week in my life.


Although I am not a particular fan of Monday mornings, this Monday morning is one of the toughest of the year! I just came back from an exciting week featuring the ISACS16 conference in Zurich and a 3-day music festival in Austria (Figure 1). It made me realize how similar festivals and conferences are. Long days, short nights, meeting new people and listening to some raw talent all day long.

Figure 1: A tough start to this week after a conference & festival double feature last week.

Figure 1: A tough start to this week after a conference & festival double feature last week.

Don’t get me wrong, I enjoyed it a lot, but it took a lot of energy out of me and getting up for the obligatory Monday morning group meeting at 9 a.m. is tough. I get coffee and arrive on time, a miracle! After the meeting, which is held in the Chemistry department, I postpone my plans to go to the Cancer Research UK (CRUK) Cambridge Institute, where the biology projects of our group happen, for one day and take some time to recover. After all, I missed a lot of science in the last week, as my RSS feed and email client tell me (Figure 2). Even better, I have to analyze some exciting sequencing data, which were generated in my absence. Having multiple, diverse projects running in parallel is one of the great things about being a chemical biologist. As my computer does all the hard work, aligning millions of reads to a reference genome, all I need to do is drink coffee and use the software correctly. The latter is something I am still struggling with (Figure 2). Nevertheless, at the end of the day I get everything analyzed and the results tell me that I am all set for writing my first manuscript as a postdoc! I also managed to catch up with my emails and the RSS feed so it’s time to cycle home and get some much-needed rest!

Figure 2: Clearly, I haven't figured out how to take screenshots on a mac…

Figure 2: Clearly, I haven’t figured out how to take screenshots on a mac…


Another morning and it is time to head to the CRUK Cambridge Institute (Figure 3), where I will spend the rest of my working week.

Figure 3: The CRUK Cambridge institute and yes, we do have the occasional sunshine here in the UK.

Figure 3: The CRUK Cambridge institute and yes, we do have the occasional sunshine here in the UK.

I am not sure how many of you fellow chemists have ever set food in a hardcore biology working environment, so let me give you a short tour. Things here are a lot cleaner and the number of fume hoods is sadly kept to a bare minimum. They are mostly used for “dangerous” phenol-chloroform extractions of nucleic acids. Being thrown into a new working environment, I always look out for things I recognize or can relate to. Lab coats are mandatory and even wearing eye protection is reinforced (Figure 4). You can also spot the occasional TLC chamber (everybody loves TLC chambers), although they are often used for a completely different purpose than analyzing your reactions.

Figure 4: Familiar sights for a chemist in a biology lab.

Figure 4: Familiar sights for a chemist in a biology lab.

Once you are feeling more comfortable in the world of biology, you might even find more similarities to your familiar chemistry lab. For working with tissue cultures, we have special hoods that remind me a lot of glove boxes. Instead of using an airlock you are using ethanol to decontaminate everything before placing it inside the hood. Of course, once you put your thoroughly washed hands inside, your nose starts to itch. Another similarity to the familiar glove box; you have to keep the place spotless as contamination with evil bacteria or yeast will spoil not only your cells, but could affect the cultures of a whole lot of other people (Figure 5). This scenario is especially bothersome, when you have worked for months creating a cell line for a particular disease you’re studying, only to find it contaminated and yourself right back at the start of your project.

Figure 5: Things you do not want find in your mammalian cell cultures! (

Figure 5: Things you do not want find in your mammalian cell cultures! (

Unfortunately for you, I won’t culture any cells this week, so I can’t show of my recently acquired and still embarrassingly clumsy skills, but I encourage anyone who’s curious to give it a go. It is surprisingly simple to culture mammalian cells like HeLa or HEK293 and as a chemist you have enough skills in your repertoire to learn it quickly. Because you have to work carefully and be gentle with the cells, I always picture myself handling tert-BuLi, which freaks me out, but my cells seem to appreciate the gentle treatment.

The plan for the week is to continue with a project I stopped working on before my week abroad. To cut a rather long story short, we identified some potential protein targets in a screen and are now keen on validating these hits. To get an independent confirmation, we need to clone all  28 proteins of interest (P.O.I.) into a transfection vector and express them inside the cell as a tagged version, in order to confirm the interaction by Western blot. That should suffice to give you a rough idea, and the rest of my day is spent planning everything and diluting 56 primers to the right concentrations. By the time night falls, my pipetting thumb has had a good workout!


I spent my PhD in an enzyme-engineering lab, so I did my fair share of cloning and from my experience I can tell you everything starts approximately like this:

Figure 6: Every good cloning starts with a successful PCR. The tricky thing is where to go from there.

Figure 6: Every good cloning starts with a successful PCR. The tricky thing is where to go from there.

For the current task at hand it is a bit trickier. For half of our P.O.I.s we were lucky and could obtain the cDNA – that is the complementary DNA synthesized from the corresponding messenger RNA – in the form of E. coli glycerol stocks that carry a vector containing the cDNA. For these proteins, cloning is easy: isolate the plasmid from the E. coli precultures and simply amplify the cDNA with the correct primers. We use primers that have 5’ and 3’ overhangs, which allow us to subclone the amplified cDNAs into the Gateway cloning system (Figure 7,

Figure 7: Step-by-step workflow of Gateway cloning.

Figure 7: Step-by-step workflow of Gateway cloning.

This method is neat, because it uses a recombinase instead of restriction enzymes and the main objective is to bring your insert into the entry vector for the Gateway system. From there, you can use another recombinase and insert your cDNA into a whole bunch of different vectors that carry appropriate tags and also allows transfecting mammalian cells! Compared to 10 years ago, when I first tackled a cloning problem, this protocol is a piece of cake.

As I started the E. coli precultures from the glycerol stocks before I left yesterday, my day consists of isolating the plasmid, doing the PCR reactions, and purifying the inserts. Sounds like a walk in the park, but it takes time (a lot of pipetting again). By the end of the day, I got 12 out of 28 cDNAs ready to insert them into the entry vector. It was clearly a successful day and I leave the lab happy for my weekly basketball game!


Today, I start the cloning of the P.O.I.s we couldn’t obtain from the cDNA in the convenient E. coli glycerol stock form. This cloning is a lot trickier, as we have to prepare our own cDNA. For this, we isolate the total RNA from HeLA cells, which involves a phenol-chloroform extraction (so dangerous!) in a real hood (so happy!). Next, we use a poly dT primer that – at least in theory – is expected to hybridize with all mRNAs in the cell as they carry a complementary poly A tail. Creating an RNA-DNA duplex allows us to reverse transcribe the whole transcriptome and generate our sought-after cDNAs. In principle, we should have our P.O.I. cDNAs in there as well, however there is no easy way of determining the concentration and whether they were fully reverse transcribed in the first place. Nevertheless, we take the crude reaction as a template for some PCR reactions. Given that we are working with a complex mixture, I start a gradient PCR – which probes annealing temperatures between 50 and 70 degrees – and hope for the best. A few hours later, I load the first 48 out of 96 PCR reactions onto an agarose gel (Figure 8), and after size separation take a look at the gel under UV light! Hurray, for 4 of 6 targets we amplified something (Figure 8). I check, whether they have the right size, which they all do, and purify them. The second batch looks equally good, which means that combined with the inserts I amplified yesterday, I got 25 out of 28 constructs ready for the recombination reaction. That’s pretty awesome and I call it a day!

Figure 8: Loading of my agarose gel on the left (hoping), results on the right (celebrating).

Figure 8: Loading of my agarose gel on the left (hoping), results on the right (celebrating).



It’s Friday! And it’s a special Friday, as we are invited to our bosses place for a British “summer” BBQ. I prepared some salmon-spinach roles yesterday night, but they really looked ugly so I didn’t dare taking a picture. With the BBQ coming up later in the afternoon, it will also be a short day in the lab, which suits me, as all I need to do is finishing the first stage of my cloning efforts.

To insert my cDNAs into the Gateway entry vector, all I need to do is mix the vector with my PCR products. I then add the recombinase enzyme that swaps the standard insert – a gene encoding for a toxic protein that prevents growth of false positives – with my cDNAs. After incubating the reaction for an hour at room temperature, I thaw some chemically competent E. coli cells, which I will use for the upcoming transformation. These bacteria are suspended in a buffer-DMSO mixture, conditions that promote plasmid uptake through the cell membrane when heated to 42 degrees Celsius for a short time (about a minute). This procedure is a bit cruel as it kills most of the bacteria; after all they don’t like the DMSO too much. However, some bacteria that took up the plasmid during the heat shock survive and they are allowed to recover at 37 degrees Celsius in a rich medium for half an hour. Next, they are pelleted by centrifugation and resuspended in 100 μL of medium. The E. coli suspension is finally spread onto LB agar plates that have the right antibiotic (kanamycin) in them, which ensures the plasmid is amplified while the bacteria happily divide. Normally, you would incubate at 37 degrees Celsius overnight, which will give you good-sized colonies; however, with the BBQ and the weekend coming up, I just place them on my bench, where they will incubate at room temperature over the weekend, delaying the growth by about two days (Figure 9).

Figure 9: My prize after a week of cloning! Grow E. coli, grow!

Figure 9: My prize after a week of cloning! Grow E. coli, grow!

All there’s left to do: Head home, get the ugly salmon-spinach roles out of the fridge, cycle to my bosses place and enjoy a burger and some beer at the BBQ. Obviously, rain begins to fall as soon as the BBQ starts, but hey, that’s life in the UK after all.

 Figure 10: A proper British “Summer” BBQ in the rain.

Figure 10: A proper British “Summer” BBQ in the rain.

Author biography:

ClemensClemens Mayer is a postdoc, working at the University of Cambridge under the supervision of Prof. Shankar Balasubramanian. He was born in Graz, where he completed his undergrad in 2008. Subsequently, he moved to Zurich to pursue his Ph.D. in the field of enzyme engineering. In 2014 he joined the University of Cambridge, where he is currently investigating the role of RNA structures in biological processes. Passions include coffee, basketball, the accumulation of useless knowledge, being a geek, and dreading the English summer.

If you are interested in writing a guest post for #RealTimeChemInFocus, please get in touch with @RealTimeChem on Twitter.