The idea appears to not be totally original, but since Tsein suddenly found fame and fortune making shit glow, I wanted to grab a slice of that pie for myself by marketing glowing shit to kids. Glowing pets aren’t new and you can buy glowing Zebra fish at pet stores and, if you wish really, really hard, you can maybe get a glowing kitteh or bunneh. Drawing on my previous experience as a child I realized that children like
- To put shit in their mouths
- Yogurt filled with sugar
- Glowing things
- Spending their parents’ money
Yogurt is even easier since it already comes preloaded with cultured Lactobacillus, a lactose loving bacteria that makes milk into tasty yogurt. The fact that it’s lactose loving and essentially just lives in the yogurt would make it an ideal candidate for transfection with a plasmid that encodes for green fluorescent protein (it’s safe) and Aequorin. Little did I know, however, that the process was a little more complicated than that.
Plasmids, it turns out, ARE NOT permanent parts of the bacterial genome and the only way to keep them being expressed in the genome is to give the bacteria an ‘offer they cannot refuse.’ I.E. attach some shit that encodes for Amoxicillin resistance and grow them in a yogurt culture with Amoxicillin delicately blended in. This way, if they don’t express the plasmid they die, and since they’re expressing the plasmid they might as well encode for the GFP while they’re at it.
This sort of left me and my charter company, Sports Milk Inc., with a rather difficult choice. Do I put Amoxicillin resistant bacteria in children’s yogurt? Why not? And while this may have sat well with me, I can just imaging all the pissed off people at “regulatory” agencies that would find this “bad” or “irresponsible.” Whateves. That means I have to actually insert the gene INTO the bacteria’s DNA. Alternately Sports Milk Inc. could market antibiotic yogurt to those crazy fucking soccer moms that have Purell dispensers attached to the outside of their minivans.
In the end, transfection with a plasmid is likely not going to be an effective strategy.
The second hurdle is actually making GLOWING yogurt and not fluorescent yogurt. Given the associated costs of producing these things, each cup will probably have to sell for about $16,000 so, you can imagine, when Junior opens the lid HE HAD BETTER FUCKING SEE GLOWING YOGURT. Sadly, the Lactobacillus used in yogurt production is a facultative anerobic organism, which means that Junior will most likely have to furiously whip air into the yogurt before it starts to glow so the Aequorin can get a good source of oxygen. That’s not a bad thing per se but kids don’t cleanly but furiously stir shit. Not only that, incorporating the GFP into the bacterial genome is going to be hard enough – also incorporating aequorin is going to be especially painful. THUS, exogenous materials must be used, for which I have a solution:
Coat granola in Cyalume and Fluorescein and include it in one of those lid-sachet thingiees with a small plastic ampule of 30% hydrogen peroxide that Junior would break into the yogurt and lightly mix. PRESTO! Glogurt for cheap.
Now give me my fucking Nobel Prize. I must now painfully do titration experiments. If you have a great program that will calculate the Ka of shit in a 1:1 or 2:1 binding mode, don’t hold back! I really want someone to tell me there’s a program that will do that for me. I poop on Origin since they didn’t bother to put any nonlinear fits that do that.



I really want someone to tell me there’s a program that will do that for me. I poop on Origin since they didn’t bother to put any nonlinear fits that do that.
Your kidding, right? Are you using a full version?
Origin is what I did all the calculations for my thesis with…. it has wonderful non-linear regression tools. I was doing 1:1, 2:1, 3:1 and 4:1 binding, with both isolated and cooperative (positive and negative)binding modes in Origin.
It is very easy.
You might try graphpad. Graphpad is not as fancy as Origin, but it does have a good UI.
If worst comes to worst, use excel and the Hill equation.
Alright. If you’re going to make me feel stupid, you have to at least spare me some dignity and tell me what I’m not seeing. I’m using OriginPro 8 (I assume the full version) and going to non-linear fit.
Are you saying there is, built into Origin, a fitting (or whatever the fuck you wish to call it) precompiled to determine binding constants? Or are you saying I can create one. Because I know I can create one. I don’t want to. I want to plug in numbers and select something from a drop down menu and have it fit to whatever binding mode my whim wishes.
A single site (1:1) binding is modeled by a hyperbola… just look for that one. I forget which variable is Ka (or Kd)… for 1:2 things get a little more complicated. If Ka1 and Ka2 are different and independent (don’t affect each other), use two hyperbolas, added together (Ka-apparent = Ka1 + Ka2). If they do affect one another, then you need at least a hill plot… which I think origin has.
Look at http://graphpad.com/www/nonling1.htm, this should give you all you need.
Remember, non-linear fitting is just applying a model to your data. The more relevant the model, the better the fit. Very few binding systems behave ideally, I have been mislead many times by this.
Yes. This graphpad is
More my speed but it still doesn’t take my tittation data and spit out a Ka in M-1. This makes me sad.
you better package the yogurt with a free UV flashlight, because GFP nor fluorescein glows in the dark. maybe you could use luciferinase?
That is what the aequorin and the cyalume is for. I am an expert on glowing stuff.
of course!
Hrhr Glogurt :>
Talked about things to attach on GFP a lot in the lab the last few days but never came on that idea. n1!
Concerning the binding modes: Never saw equations for 2:1 somewhere in literature. So if it’s possible with Origin’s build-in functions, I would be highly surprised. Maybe there are some binding models integrated, which then apply to proteins only IMHO…
You’ll find “Specfit/32″ sometimes in lit for complicated binding modes, cause it has a very mathematical approach to analyze spectra. I think there’s a limited demo free of charge available. But be warned, it’s very complex and I never came that far to understand what’s the program actually doing…
“But it comes with a free glogurt!”
“That’s good!”
“The glogurt is also cursed.”
“That’s bad.”
“But you get your choice of topping!”
“That’s good!”
“The toppings contain potassium benzoate.”
“…”
“That’s bad.”
“Can I go now?”
Awesome, thank you. Just watched that episode on Saturday.
This post, sir… is why you are a great blogger.
thks!
You’ve commited a trademark violation.
The Dannon gestapo will hunt you down. I once made a joke about the Jolly Green giant on my blog and I have been on the run ever since.
Biochemistry is molecules and crap bothered by somebody too lame to do the work himself. Yogurt plus microencapsulated tetrakis(dimethylamino)ethylene; chew. Call it GlowMouth(tm) yogurt. It will satisfy kids’ craving for crazy chlorine bleach flavor, too.
Actually, maybe you could put the GFP code into an adenovirus, and presto, glowing kids!
Instead of using antibiotics, the smart ™ way to do this is to use a Lacto auxotroph, that is incapable of synthesizing, say, serine. Then put the serine synthase on the plasmid and you’re good to go.
From a parent…glowing food inevitably (or maybe not?) leads to glowing poop…I can only imagine where this leads to. What’s the effect of pH on the suggested active ingredients?
glowing poop leads to kids running to their parents saying “LOOK AT MY POOP!” which I have approximately zero problems with.
This is the best! Glow in the dark cat!