Sometimes it feels a little bit too good to spend money in the lab.
Blue streak If you have to ask, you can’t afford it
For those of you following the story so far, I recently triumphed in my new career as a Cloner For Hire. In brief, I managed to make and verify three complex DNA constructs, finishing the very day my visa-frazzled colleague returned to London to pack for his sabbatical. This had already turned out to be a bit more involved than I had initially anticipated – and then the boss suggested I extend the favor by recombining the little blighters into baculovirus vectors (so-called ‘bacmids’) so that my colleague could furiously finish up a few final experiments for the paper in the week before his departure.
The bacmid kit seemed straightforward, but I did notice that the transformation step – the part of the procedure when you zap bacterial plasmids into E. coli and allow them to recombine with a resident bit of DNA – looked a bit problematic. I could tell something was up by the cagy way that the kit’s instruction manual phrased certain things. That the procedure was far from efficient was also clear by the modifications in the standard protocol: five hours of growth before plating onto the triple antibiotics plates instead of thirty minutes; two days’ growth after plating instead of one; and then a further re-streak of ten prospective white colonies to confirm that they actually were bona fide recombinants.
At first I thought the company was just trying it on when they suggested purchasing their own special – and highly expensive – derivative of X-gal, the chemical substrate you need to put into your agar plates so that you can tell which bacterial colonies have successfully recombined (blue colonies are unperturbed, whereas white colonies are bacteria that have lost the beta-galactosidase gene because the genetic modification of interest has disrupted it). But then I thought about it: two days to get color development and a re-streak to confirm it? These colonies were likely to be pretty anemic, and even freshly prepared X-gal can often result in very pale colonies. We simply didn’t have a day to lose on ambiguity.
Dear reader, I bought that expensive X-gal derivative. And I have never seen such lovely, deep-blue, almost slate-purple colonies in my life. I thought I would weep with joy, and my colleague as well – it was KimWipes all around. And it’s a good thing I did too, because after re-streaking the white colonies, the blue controls were so faint the next morning that I spent many, many minutes squinting at the colonies, trying to decide if there was the faintest whiff of blue about their whiteness.
The moment of truth came in the form of PCR confirmation. There is a wonderful moment in science just before a result, no matter how routine. For me, even after two decades in the lab, I still get a frisson of anticipation when an agarose gel is sitting on the lightbox, the verdict unclear in that hushed moment just before the UV source is switched on:
And then –
– you see that your bands of DNA spell out success. It’s a priceless feeling.
Just an hour before the Fedex deadline, I was coaxing my final bacmids into solution and labeling up the tubes to ship them off to my colleague’s sabbatical lab; with a prevailing wind, they’ll rock up at about the same time tomorrow morning as he does.
Time, I think, to saddle up and head off into the sunset.
five hours of growth before plating onto the triple antibiotics plates instead of thirty minutes; two days’ growth after plating instead of one.
yup, I recognize that idea…. triple antibiotics make everything slow and sometimes weird. My bacteria tend to be a bit sad about their capsule when more than one antibiotic is in the plate.
It’s fun that you got it to work!! Good luck with the shipping! And I agree with you about turning the gel on. the anticipation 🙂
\o/
Asa, I often find myself getting a bit sad about my own capsule after all these years — I don’t even need the antibiotics.
There is a wonderful moment in science just before a result, no matter how routine.
It’s always going to be like that? I was thinking I’d grow up eventually…
I can only speak for myself, Anna! But it’s always been like that for me — the darkroom is the scene of most of my mini-dramas, but even looking down the light microscope at my cell cultures gives me a good feeling.
{valiantly resists fnar-ing}
Anna. You have to grow old, but no one has to grow up.
That’s my philosophy, and I’m sticking to it.
If the joys of science are gone, as far as I’m concerned it’s time to pack it in. Because otherwise all you’ve got left is the despair.
Never a truer word spoken.
At least I was still enjoying it when I packed it in. So memories are more happy than sad.
And they’ll just get happier, if my experience is anything to go by…
Well, if they get too happy, I’ll come in at the weekend and do some cloning or set up crystal screens for you, or summat.
Split your cells, ma’am?
Don’t tempt me.
I’m serious.
In other news…it turns out I am actually faster and more reliable than Fedex!
Ok you two, get a
darkroom.It really is wonderful when something clones nicely, isn’t it? I remember that feeling well, long ago though it was. I used to get the same rush from a significant p-value too. When the data hand you the result, everything seems good.
Oh, and If the joys of science are gone, as far as I’m concerned it’s time to pack it in. Because otherwise all you’ve got left is the despair. is going to be my new .sig over at this place, if that’s alright with you. Properly acknowledged, of course. 😉
Of course it’s all right! I’m flattered.
I’m afraid my joy in science doesn’t extend to favorable statistics — probably because I’ve never quite managed to get my head round them.
Oh, don’t get me wrong, I don’t understand the stats – just enjoy them when they spit out something that looks like a significant result.
.sig updated. 🙂
That was a thorough answer! It’s comforting to know that the excitement won’t go away – but I still wish I didn’t get quite so nervous. Waiting for the film to go through the developing machine at the end of an important experiment (those minutes are SO long) still makes me feel slightly sick. Maybe I don’t have the nerves for this job 😉
I think the nervousness is an integral part of the fun. If you didn’t care about the outcome, it wouldn’t be worth anything.
I only get slightly sick if it’s an exceptionally important experiment. 🙂
I used to say that the moment I stopped being thrilled by science would be the moment I knew it was time to stop doing it.
But it didn’t quite work out like that.
Psst. He didn’t jump — he was pushed. Always suspected foul play.
Cluck.
Nice, though my favourite is S-gal, which gives black colonies – no missing those blighters!…and you can autoclave it.
I thought I would have a heart attack the first time I tried a trial pack of the mysterious ‘Harlequin agar’ I was offered as an alternative to X-gal quite a few years back – all that time wasted ‘developing’ the blue, only to get blue, off-blue, off-white and white colonies. Never again.
p.s. “We simply didn’t have a day to lose on ambiguity. – fantastic.
Jennifer and Richard,
You are right of course – it’s much preferable to be thrilled on the verge of feeling sick about ones work than to be bored into the “I couldn’t care less” state. I have absorbed your wisdom and will hereafter enjoy any “pre-result” nausea or dizziness to the fullest 🙂
Yay!
I like the thought of evil black colonies, raging an eternal battle with the white good colonies.