Some of my friends spent their long August bank holiday weekend in the lab. (Luckily, nobody from my lab – I was in on Tuesday and the place was dead even then!) Some of them took the opportunity to have a bit of a holiday. Me? I went to BABS Harmony College.
No, really. Think “band camp” but for barbershop. Yes, it’s a real thing. Yes, I am aware this sounds like the least cool thing ever – and I’m a scientist. I wasn’t cool to begin with. As it happens, it was just about the most fun I’ve had in the whole of 2012. (Yes, more fun than an impromptu 2 weeks in Copenhagen for work. Yes, more fun than learning twisting somersaults. Trust me, I really enjoyed this).
Anyway, this is a biology blog right – so why am I talking about barbershop?
It is infuriating to discover you have slipped up somewhere and ended up with next to no DNA or RNA or whatever at the end of your prep.
It is even more frustrating when you had planned really well and done everything to the letter.
But the most frustrating thing of all must be to be certain you have done it all right, have it all go wrong anyway and then discover someone else in the lab already knows why.
I have (had? *sadface*) some RNA. I DNA-ase treated it prior to using it in qPCR. It needed cleaning up.
In the previous incarnation of this experiment I cleaned up the RNA (Qiagen RNA MinElute) then DNA-ase treated it. (Turbo DNAfree, in case you’re interested).
Except that the more I thought about it the more I worried that I was leaving residual DNase behind. So this time around I used the Turbo DNA-ase and then cleaned it up using an RNease Mini Kit. Exactly like it says you should. And all of my beautiful RNA is… well not gone. But coming out at ~50ng / uL. FIFTY. What the actual hell?! The other way around I got nearer TWO THOUSAND!!
Now obviously a MinElute elutes into 14 uL not 30, so it’s only natural to expect the prep to be half as concentrated: let’s say it was a pretty rough prep that would have had 800, then I might expect 400. But fifty?! Absolutely gutted and so unbelievably cross with myself for throwing all 14 samples at it at once.
But the most annoying part of all? Another PhD student in the lab figured this out about 2 months ago. Turbo DNA free and Qiagen RNeasy mini just aren’t compatible with one another and as far as I can see nobody has ever made a note of this.
Why do we always want what we can’t have? Over a couple of months of furious labwork I successfully blogged at least every other day. While waiting over a week for some Brilliant III to surface so I could crack … Continue reading
There is a school of thought, as championed by Simon Baron-Cohen, that thinks of autistic disorders as ‘an exaggerated version of maleness’. The theory goes that autists are systematic, unable to articulate feelings, not great at empathising. They tend to gravitate towards logical subjects like maths and science. They often don’t have close friends. The same things are broadly more true of men than of women. Female diagnoses of high-functioning autism are much rarer (4:1 for profound autism, maybe even 9:1 for Aspergers). Ergo, maybe autism = extreme maleness. Girls are less likely to be diagnosed because a particularly “male” girl is… well, a tomboy. Whereas an extra-“male” boy is more of an anomaly.
It’s a nice theory, but has not been conclusively shown to have a physiological or endocrinological basis. The empathising / systemising nature of humans in general has been shown to correlate well with testosterone levels in foetuses. Testosterone has been shown in vitro (i.e. in a petri dish) to have an inhibitory effect upon a transcription factor called RORA, and biopsies show that autists have lower levels of RORA in their brains than the general population. There could very well be a link, but it’s not conclusive. It also doesn’t tell us much about why there is a link.
Why are you in work so late?! asked my housemate’s boyfriend, when I explained via Facebook chat that that was where I was.
I got really into writing. And now I’ve been here so long that the qPCR I planned to run over night is done, so I may as well do the analysis! I replied
Dedication to the cause! Came the reply. But I have no idea what qPCR is…
I pride myself on talking about science far too much and boring everyone around me, especially housemates and their boyfriends. So how have I possibly avoided explaining what qPCR is?!
DNA exists in relatively small quantities compared to how much we need to do molecular biology. In order to work with it – or, for that matter, check whether it is there – we need to make more of it. A particularly awesome feature of DNA makes this possible. It is a double stranded molecule: if you could straighten out the helix that it works itself into it would look like a ladder, and the two sides are inverse copies of one another. (Once you have finished marvelling open-mouthed at my non-existent artistic skills you will, I’m sure, spot that pink always pairs with orange etc).
This means that if you split the molecule in two, it’s possible to rebuild the other side from the side that you have. This happens inside your cells all the time and is called semi-conservative replication.
The Polymerase Chain Reaction
We can simulate this in the lab through a reaction called PCR (Polymerase Chain Reaction). PCR can best be summed up by my favourite geeky advert of all time. (A geeky advert so immense that for my first 4 months as a PhD student I drank from a BioRad mug for no other reason).
In case that was all a bit much for you, here’s a quick recap:
I’m sure I’m not alone in having regularly received lab supplies – especially enzymes – shipped in bizarrely large boxes. Today’s offering, however, really takes the biscuit:
This box (with autoclave tape for scale) contained four of these tubes (with my hand to scale). In fairness, it was shipped over the weekend, but all the same that is a shedload of dried ice for four very small tubes!
Amusingly, today I also received this.
This is also a qPCR enzyme and also says that it should be stored at -20C. This one however was shipped in a plastic bag with a (completely melted) ice block. Fingers crossed it still works… not planning on using any precious samples to test it though!
Tonight has been a busy night. One of the PhD students in my office, V, has her viva tomorrow. I will be shocked if she doesn’t pass with flying colours, but will be crossing my fingers for her anyway! V loves all thinks pink and flowery, so as Chief Cake Baker of the Genomics group I have been tasked with providing large quantities of pink cake. Continue reading