creative commons

Life is, as ever these days, quite busy! I have two trips coming up, one to Ignite in Dublin and one to the Newcastle Maker Faire (where Brian Degger of Transitlab.org and I will be hosting a little workshop on a few DIYbio experiments and fun stuff). In both cases, I have to have stuff prepared and ready to demo, and in both cases I’m tentatively on-schedule. Which is nice for a change.

For the latter event, I’ll be giving a presentation on how to isolate and culture bioluminescent bacteria from fresh seafood, which people following my twitter feed will have seen me gibbering about for a while now. The first step in culturing any microbe, of course, is cooking up and sterilising an appropriate growth medium in the form of broths and agar dishes/slants.

In order to cover that base, I created a protocol to cook up lab-quality growth media using only off-the-shelf ingredients, most or all of which should be available at a pharmacy and/or health store. The only ingredient not available in ready-to-use form is the peptone that most media call for; I include a recipe and super-easy instructions on how to make peptone from Soy (called Phytone) or Casein (similar to Tryptone) using the enzyme Bromelain, which is widely available as a digestive aid or as meat tenderiser and performs quite well at a variety of temperatures and pH conditions.

Without further timewasting, here is the PDF (~7MB) and the OpenOffice.org Document (~10MB). This work is released under an Unported Creative Commons by-sa license. Apologies for the size, there’s a set of images embedded. Here’s a PDF and Document lacking the images, much tinier and easier to load on phones.

I can comfortably endorse the use of these media: Using an LB batch I brewed up using this method (and using Calcium Carbonate antacid tablets instead of Sodium Hydroxide), I grew up a batch of E.coli bearing a plasmid I needed in the lab. Using the Luminescent Broth as described, I’ve isolated bioluminescent bacteria and found to my delight that they grow and glow green-blue stably for days on an agar stab or on media wicked into a piece of sterile tissue/cloth. They grow in the medium on its own, but won’t glow if there isn’t enough surface area for gas exchange, leading to dark mutants taking over the medium. I’d share pictures of my successful strains glowing happily, but I don’t have a camera that will do long exposures.

I’ll soon be writing up a companion document telling you how to *use* your media to isolate bioluminescent bacteria, but that protocol is more or less identical to those detailed already on the Indiana Biolab disknet archive and on Mac Cowell’s blog. I’ll only add some practical details for beginners on how to streak agar plates, which can really help you to separate your brightly glowing bacteria from contaminating strains and dark mutants.

More on this soon, and if you perform anything inspired by this stuff please let me know! As always, exercise caution and wash your hands all the time. Right now, perhaps?