Colony: Blind Spot
January 25, 2017 10:40 AM - Season 1, Episode 4 - Subscribe

Will finds the source of the radio broadcasts. Katie deals with a staged firebombing to get intel. Phyllis reveals a depth of knowledge. Maddie figures out an angle for her insulin supply.
posted by nubs (11 comments total)
 
Not a bad episode, but one I found a little frustrating at times...the Resistance expecting that the firebomb would lead to Katie getting intel is one thing, but expecting it to mean a chance to identify Phyllis seemed like a stretch. Of course, so did the interest Phyllis took in the case until it became clear that she knew far more about Katie than Katie suspected.

While I figured Phyllis was on her way out last episode, the show did it really well - she was a interesting character, and I thought she added some nice complexity to the situation - not just in terms of knowing about Katie, but in her frankness about the situation; that everyone is lying about what is happening, and that she views her role as trying to stop the Resistance not because she believes in the Raps but because she believes that the vengeance the Raps would take for Resistance activities would be massive and unstoppable. Kinda sorry to see her go, but it made sense storywise...now, where is her file on Katie?

And Broussard is working for Homeland, which is interesting. I wonder if I go back to episode one if he's the guy who checks Will's ID in the truck...

Still struggling to care about Maddie's plotline. It feels tacked on and that we are supposed to care because she has a sick kid. But interesting to learn that the Raps are apparently interested in our art as some pieces have "gone up". Maybe the Raps are just the 1%?

Will certainly seems to have the biggest blind spot of the moment, with not seeing what Bram and Katie are doing. But everyone seems to have a bit of a blindspot going on...including the fact that "Geronimo" is apparently in the Green Zone where the most trusted people live.
posted by nubs at 10:55 AM on January 25, 2017 [1 favorite]


Kinda sorry to see her go, but it made sense storywise...

I know! I really loved Phyllis as a character, and the actress as well. So bummed we're not gonna see her again.

I wonder if I go back to episode one if he's the guy who checks Will's ID in the truck...

Heh. That would be a totally unnecessary coincidence, but it'd be neat if so.

Still struggling to care about Maddie's plotline. It feels tacked on and that we are supposed to care because she has a sick kid. But interesting to learn that the Raps are apparently interested in our art as some pieces have "gone up". Maybe the Raps are just the 1%?

This is one of the problems with a slow-burn show. Sometimes, you have a subplot that you want to bring in towards the end, but that only works if you cut to it throughout the season to keep it present in viewers' minds, even if there's not much to actually show.

The idea that the Raps might be humans who live in some kind of space habitat is interesting. Their tech is advanced, but nothing we've seen so far is completely alien. The drones are something I could easily see humans making within a few years of our present day.

But it'd also be pretty interesting if they did turn out to be aliens that just happen to have an appreciation for art and culture, even if that appreciation is more along the lines of cataloguing than enjoyment.
posted by tobascodagama at 12:13 PM on January 25, 2017 [1 favorite]


This is one of the problems with a slow-burn show. Sometimes, you have a subplot that you want to bring in towards the end, but that only works if you cut to it throughout the season to keep it present in viewers' minds, even if there's not much to actually show.

Yeah, I get it. I mean, there's totally a sense that what Maddie is doing is going to somehow lead to something important somewhere, somewhen, but it's hard when every other storyline actually seems to be advancing, while she's out trolling for insulin.
posted by nubs at 12:49 PM on January 25, 2017 [1 favorite]


The art subplot is fascinating for me, especially as it has - intentionally, I suspect - resonances of the kind of stuff that went on in Europe during WWII in Nazi-occupied countries.
posted by Rock Steady at 5:34 AM on January 26, 2017


Enjoyed the COPS opening, and glad to have media address toxic authority-inspired force-escalation.

I thought that the insulin subplot is interesting; looked briefly for the original insulin-cures-diabetes-in-people papers, and found them, but hadn't yet found the original with the methods on how to extract insulin from dogs. The original paper referenced it as "to be published later." They did give a brief overview and it's pretty crude - but apparently worked. (Ethanolic extract of dog pancreas, evap, wash, resuspend in saline - then a unit assay to determine potency, of which a 1922 paper detailed). Surprisingly (Not surprisingly), canine, porcine, and bovine insulins all work in humans almost as good as the human version.

Gonna spend some time tomorrow (or maybe next week, my weekend started a few hours ago) looking up the most practical way of making some homebrew insulin. Recombinant yeast can be created with a 'minimum' of (molecular biology) lab equipment this is predicated on continued access to the electronic literature (importantly, genbank).; fermentation and purification should a breeze given some 'basic' glassware. It for sure won't pass the FDA, but there are low-er tech assays that tests if something is non-harmful [for a given limited set of harm] to inject into someone.
posted by porpoise at 6:39 PM on January 26, 2017 [2 favorites]


Aliens and art: Theft of cultural artifacts was par for course during imperial colonialism. Most of the invaders viewed the conquered as a lesser. A lot of the art was of a completely different tradition and would have seemed alien to the conquerors. Sometimes, different values were attributed to materials and the technology used to shape those materials (cast/worked gold vs carved stone or wood) but the prized art of the subjugated was still appreciated by the colonialists.

I easily could see aliens looting cultural artifacts that Earthlings thought was valuable, even if merely as a show of power (qv the desire, drive, and insistance for the repatriation of ancient Chinese artifacts, back to China, that were looted over decades).

> "Geronimo" is apparently in the Green Zone

So the guy they captured... memorized realGeronimo's speeches or is just a freelance mouthpiece? Or is the resistance organizer just taking advantage of a freelance pirate radio broadcaster?
posted by porpoise at 6:43 PM on January 26, 2017 [2 favorites]


Your questions are answered next episode. :)
posted by tobascodagama at 7:36 PM on January 26, 2017 [1 favorite]


Gonna spend some time tomorrow (or maybe next week, my weekend started a few hours ago) looking up the most practical way of making some homebrew insulin. Recombinant yeast can be created with a 'minimum' of (molecular biology) lab equipment this is predicated on continued access to the electronic literature (importantly, genbank).; fermentation and purification should a breeze given some 'basic' glassware. It for sure won't pass the FDA, but there are low-er tech assays that tests if something is non-harmful [for a given limited set of harm] to inject into someone.

I would be very interested in hearing about that.
posted by Rock Steady at 5:32 AM on January 27, 2017 [1 favorite]


Without teaching the last two undergrad years of molecular biology (or maybe, this is closer to a MSc)...

There are a number of integrative plasmids in yeast that will incorporate into their chromosomes. Plasmids typically have a selection cassette already integrated. There are a number 'tamed' yeast strains. Yeast is important since they are eukaryotes - mammalian insulin is more likely to fold correctly in them than in the easier to manipulate/engineer e.coli and you don't have to worry about endotoxin.

Insert the full length cDNA (the complementary, complete, spliced sequence of the coding mRNA) of human insulin into the plasmid. This can be obtained either as fully synthetic or you can clone it from a human (or whatever) cDNA library (which can be constructed de novo).

Engineer in a 1) secretion leader peptide and 2) an affinity peptide for chromatographic separation. The choice of which one is tricky since the effectiveness of the different types will vary and it probably has already been determined. But something like biotin/avidin or IgG/proteinA/G. Which to use will probably depend on whether you have access to biotinylated beads or proteinA(or G) beads or whichever one you can make from scratch.

Engineer in a 3) cleavage site, again, the specific choice of cleavage site depends on what else you have on hand. If you have the technical capability to get to the endpoint here, its trivial to clone, express, and purify any number of cleavage enzymes. Again, the efficiency of the different cleavage sites has probably already been determined.

Once you have the engineered plasmid with the recombinant human insulin, you transfect that into yeast and grow them up a little. Plate them on selective media so they expand into clonal colonies. The ones that survive/thrive on the selective media are the ones that have the plasmid integrated into them. You select a few and screen them for activity. If you have an antibody to hInsulin, great - you can screen by Western blot and select the one with the highest activity. Trick here is that chromosomal integration is semi-random (but there are tricks that you can use). Depending on where it integrates, the activity of the engineered gene may differ, or may interfere with the yeast's normal functioning.

If not, then its more laborious and you go in blind and screen extracts for activity after fermentation.

Anyway, once you have a stable line that you like, you grow them up in liquid culture with... essentially... dead yeast soup. The engineered yeast will make rhInsulin and secrete it into the liquid media.

After a while, you harvest the media (either just take the excreted rhInsulin or lyse the yeast - which version is more efficient depends on how good your secretion signal is, and what affinity peptide you use and how you pack your chromatographic column). You'll want to "clean it up" by centrifugation and/or filtration. How 'clean' you can get it depends on what you have on hand.

Prepare a affinity chromatography column; basically a vertical tube packed with (agarose, usually) beads with your paired affinity peptide. Run the media through the column, perhaps more than once. The engineered rhInsulin will get stuck in the column and you wash everything else away. Then you introduce the cleavage enzyme/agent (which is typically innocuous to the subject the rhInsulin will be adminstered to, or you get rid of it by differential precipitation), and you wash out the column and collect the cleaved rhInsulin.

Rotovap at low temperature (the machine can be made from scratch; crude centrifuges too but both of these equipments are abundant and common) or lyophilize, resuspend in saline, run it through a 0.22um filter to sterilize. UV sterilization is an option, but less efficient and reliant on having UV bulbs, which are also a consumable. Or a gamma source if you have one.

Do the unit activity assay and dilute the final product to the appropriate concentration.

Would probably do stuff like run gels and stain for total protein at different points to determine purity. Lots of 'moving parts' where optimization would have to be performed. Also a limulus amaebocyte assay for endotoxin, but if that's not available or you don't have access to horseshoe crabs, you can do the pyrogen assay in rabbits (and potentially other animals).

This could be done in any reasonably equipped undergraduate molecular biology lab, and much of it can be improvised from other common household equipment. A core piece of equipment is a decent PCR machine, but those can be salvaged if unavailable, or a very crude version can be jury rigged from household equipment. A (DNA) sequencing machine (and all the reagents needed to operate) is in practice required for quality control of your engineered DNA while you're engineering it. The enzyme reagents can be made, and crude versions using primitive chemistry can be jury rigged, but getting polyacrylamide of sufficient purity may be difficult. You do need, however, a pretty wide range of enzymes to do the DNA engineering work, and a way to synthesize oligonucleotides. But if you have an oligosynthesizer, you could theoretically bootstrap yourself there if you start out with some polymerase enzyme (which allows you to make more polymerase similar to how you're making synthetic recombinant human insulin right now). Unfortunately the chromatography may be limiting if the supply of affinity beads is limited; but with a little chemistry (and a way to make synthetic peptides/proteins as we are doing here), those can be homebrewed, too.

--

But yeah, depending on your technical capability, it might just be more straightforward to harvest insulin from dogs (as apparently in the show)... which unfortunately will, well, die from the pancreatectomy. Having access to a slaughterhouse will give you access to fresh food animal (beef, pork) pancreases - which is basically what we did before the development of molecular biology.

--

Now ask me about antibiotics =)
posted by porpoise at 10:09 AM on January 27, 2017 [4 favorites]


I'm hanging out with porpoise after the apocalypse.
posted by Rock Steady at 10:17 AM on January 27, 2017 [3 favorites]


Bring me pancreases!

(And Thymuses - they're great eating! sweatbreads!)
posted by porpoise at 7:29 PM on January 27, 2017 [3 favorites]


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