I’ve been fascinated by bioluminescence from the first time I saw pictures of deep-sea angler fish. Since then, I’ve seen dozens of live bioluminescent creatures at the Boston aquarium and cultured E. coli expressing the LuxCDABE operon from Vibrio fischeri in Tom Knight’s lab.
It surprisingly easy to grow bioluminescent microbes. I’ve found a couple of undergraduate-level protocols online for isolating marine Vibrio species from fresh squid. I’m currently in the middle of my second attempt at isolation. These are my notes so far. I’ve adapted this Basic Protocol for Isolating Bioluminescent microbes for use in my kitchen.
I’m using food-grade agar I got from a food-hacking friend, powdered chocolate jello (I ran out of powdered agar), Baking Soda instead of CaCO3 (chalk), and canned tuna water instead of powdered LB broth (hope it doesn’t have preservatives in it).
13 Jan 2010
18:30 Got fresh, “uncleaned” squid from New Deal Fish Market in Union Square: $2.50
18:55: got 1L gatorade
Added 1/2 teaspoon of sea salt to the empty gatorade bottle
22:00 cut squid head in 1/2, added to tap water in gatorade bottle
14 Jan 2010
22:00 It GLOWS! wow!
took pictures with ad-hoc cardboard camera frame; cutting gatorade bottle in half (squid now in what resembles a petri dish with 8-inch tall walls)
Added 15 mL of Jeff’s Food-Grade agar to 450 mL of water; not enough? Still liquid. (only have a 15 mL falcon tube, not a scale).
Boiling the agar liquid in a water bath. Will add Jiffy CornMeal to supplement.
22:25 agar solution at 175 C. Cools to a mildly viscous liquid.
22:30 adding 15 mL chocolate JELLO
How many grams is 15 mL of agar powder? 0.34 g / cm^3 (http://www.wolframalpha.com/input/?i=15+mL+Agar) 15 mL ~ 5g.
How about Jello? 1.1 g / cm^3 (http://www.wolframalpha.com/input/?i=15+mL+Jello) 15 mL ~ 17g
Most LB plate protocols call for 15 grams of agar powder / Liter. I started with 5g (15 mL) in 450 mL of water. So I was 2/3 short.
It turns out 15 mL Jello + 15 mL agar in 450 mL boiling water solidifies into something approximating an agar plate. I’m adding 2.5 mL more Jello to thicken it a bit.
Other ingredients of recommended for Luminescent Agar plates (per 450 mL H2O):
- 2.5 g CaCO3 (NaHCO3 is 2.173 g/cm3; and I’m going to substitute it. Added ~ 1g)
- 5 g Glycerol (left it at sprout)
- 15 g NaCl (2.165 g/cm3, added 4 mL; ~ 8.66 g)
- 4 g “dehydrated Nutrient Broth” (I’m using a teaspoon of liquid from a can of tuna)
0:00 15 Jan 2010 Poured “Luminescent Chocolate Agar”
0:30 poked bright blue bioluminescent patches on the squid carcass with a toothpick and streaked them out on the choco-agar.