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| maaku | kanzure: that's really precise imagry for activation maximization | 04:43 |
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| maaku | FourFire: I don't know anyone that has insight into what calico is doing | 06:07 |
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| kanzure | FourFire: human longevity inc is selling genome data to insurance providers. but it's not calico. | 06:21 |
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| maaku | man i wish i bought nintendo stock | 06:31 |
| chris_99 | heh | 06:32 |
| kanzure | twitchplayspokemon might be doing pokemon go soon | 06:42 |
| kanzure | i learned last night that they have been using some of my work (pokecrystal) | 06:42 |
| kanzure | koolboyman has ported his hack rom from gold to crystal | 06:43 |
| kanzure | which is pretty funny because that migration wouldn't be necessary if i had chosen gold as the base instead of crystal like everyone insisted to me :D | 06:44 |
| kanzure | i think they are planning to debut his hack rom on twitchplayspokemon as an exclusive | 06:44 |
| kanzure | in the mean time there's a let's play ... https://www.youtube.com/watch?v=HkC6t3Iwi3Q&list=ELW8zJX5NpDLk | 06:45 |
| kanzure | s/like everyone insisted to me/like everyone insisted and as i ignored | 06:49 |
| kanzure | er... well anyway, there's a way to make that sentence work somehow. | 06:49 |
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| kanzure | fenn: "Can a biologist fix a radio?" was the inspiration. it's just a trend of copying clever-sounding titles. | 06:58 |
| kanzure | re: free floating controlled polymerase, the reason why :| is because andrew wants to do electrical control of /many/ free floating polymerases in solution simultaneously and then look for DNA that matches the given electrical input...... | 06:59 |
| kanzure | andrew and ethan kurzweil saying things http://www.bloomberg.com/news/videos/2016-07-13/why-invest-in-moonshots | 07:02 |
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| fenn | these people have a weird idea of what qualifies as a "moonshot" | 07:38 |
| fenn | IP cameras? really? | 07:38 |
| kanzure | just got off the phone with russell | 07:48 |
| fenn | bah "technology to make the world radically different" and they didn't even mention AI | 07:48 |
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| kanzure | their funding is mostly grants at the moment | 07:50 |
| kanzure | need about $3M to pursue an entire library of aptamers for about ~120 brain cell surface receptors (for contrast agent imaging/labeling reasons) | 07:51 |
| kanzure | i guess he's doing the long slog of aptamer development, although there are commitments from DFJ to move forward on condition of imaging an entire mouse connectome first | 07:52 |
| fenn | dfj is funding academic labs now? | 07:52 |
| kanzure | which i think is a big requirement.... doing this by grants for funding is going to take a long time, i think. | 07:52 |
| kanzure | nah i mean they gave a commitment to fund a venture including the development work to translate from mouse to human | 07:52 |
| kanzure | but it's conditional on doing a mouse first | 07:53 |
| fenn | well that's a big milestone | 07:53 |
| fenn | image a cell first ha | 07:53 |
| kanzure | he has existing data for axon-resolution imaging from synchotron CT scanning | 07:54 |
| kanzure | he mentioned one problem is clearing out aptamers from a live brain after imaging | 07:55 |
| kanzure | i think you can just wait a long time and the aptamers should go away at some point? | 07:55 |
| fenn | they will get degraded i guess | 07:56 |
| fenn | by nucleases | 07:56 |
| kanzure | but then you end up with different scans of different receptors at different times-- which is like getting 120 different brain scans each a month apart from each other (instead of a single brain scan on the same day) | 07:56 |
| fenn | if only we had a way to print rna in a cell on demand | 07:57 |
| fenn | this all sounds like permanent vaporware but i'd love to be proven wrong | 07:57 |
| kanzure | what data would you want to see to get convinced it's not vaporware? | 07:58 |
| fenn | a slice of tissue and its connectome | 07:58 |
| kanzure | well you will only see the contrast imaging nanoparticles or whatever, not an actual connectome | 07:58 |
| fenn | sure someone has to write the image processing software | 07:59 |
| kanzure | say you have a neuron that targets something 1 cm away, | 07:59 |
| fenn | but there are a lot of steps before you get a live brain's connectome | 07:59 |
| kanzure | i don't think receptor labeling is going to show you that this particular neuron has anything to do with its target 1 cm away | 07:59 |
| fenn | i mean there's a lot of AND's chained together in serial before it will work | 08:00 |
| fenn | this has to work AND that has to work AND the other has to work ... | 08:00 |
| kanzure | you could get all the synapse weights from this but you wont be able to figure out where neurons are connecting | 08:00 |
| kanzure | am i missing something? | 08:01 |
| fenn | is there no generic neuron membrane protein they can stain for? | 08:01 |
| kanzure | your image would just be a big blob of color | 08:01 |
| fenn | no it would be a lot of blobs of color | 08:01 |
| kanzure | shadows from other neurons? it's the same reason why you can't see through a brain (most of the time). | 08:02 |
| kanzure | i guess physical scanning on multiple perpendicular dimensions is helpful for this part | 08:02 |
| fenn | there's gotta be some way to stain for something that gives you cell shape | 08:03 |
| kanzure | sure that's not a proble | 08:03 |
| kanzure | *problem | 08:03 |
| kanzure | i'm suggesting that scanning all of that data will not give you deep membrane shape data | 08:03 |
| kanzure | so basically you are asking for (1) a high-density tissue contrast agent and (2) a scanning technique that can spatially resolve deep-tissue labeled structures | 08:04 |
| fenn | yep | 08:05 |
| kanzure | i feel like people would be talking about both cell membrane labeling and synapse imaging then, because high-density cell membrane labeling and imaging is an interesting/useful tech development too. but for some reason i only hear about sub-synapse resolution? | 08:06 |
| fenn | well synapses are made of cell membranes.. | 08:06 |
| kanzure | i guess the individual cell structure of anything not brain is irrelevant and brain matter is the only interesting organ where we we think that might be relevant ? | 08:07 |
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| bluebear_ | sorry if this was already mentioned (I can't watch the video... @work) but a modified rhabdovirus (rabies) is a good agent for crawling through neural connections and can also mark them; of course you get only what connects to what but no details about receptors, so only a very rough schema | 08:07 |
| fenn | i guess people don't realize you can guess the synapse type from morphology alone and so they try to stain for neurotransmitter receptor proteins or whatever | 08:07 |
| kanzure | synapse type and neuron type inference is often done based on neuron morphology sure | 08:08 |
| fenn | the bloomberg video is just andrew hessel talking about human genome project | 08:08 |
| kanzure | the sub-synapse resolution imaging stuff is only for weights | 08:08 |
| fenn | "weights" is artificial neural network lingo, you should probably say receptor density | 08:10 |
| fenn | or is everyone crazy? | 08:10 |
| kanzure | "receptor density" is acceptable. i've said that before i think. i'll try to stick to that. | 08:11 |
| kanzure | .wik synchrotron X-ray tomographic microscopy | 08:11 |
| yoleaux | "A CT scan, also called X-ray computed tomography (X-ray CT) and computerized axial tomography scan (CAT scan), makes use of computer-processed combinations of many X-ray images taken from different angles to produce cross-sectional (tomographic) images (virtual "slices") of specific areas of a scanned object, allowing the user to see …" — https://en.wikipedia.org/wiki/Synchrotron_X-ray_tomographic_microscopy | 08:11 |
| kanzure | grr | 08:11 |
| fenn | hey the bot worked | 08:11 |
| fenn | it's just a CT scan with synchrotron as the x-ray source | 08:12 |
| kanzure | .wik nanotomography | 08:12 |
| yoleaux | "Nanotomography, much like its related modalities tomography and microtomography, uses x-rays to create cross-sections from a 3D-object that later can be used to recreate a virtual model without destroying the original model, applying Nondestructive testing. The term nano is used to indicate that the pixel sizes of the cross-sections are …" — https://en.wikipedia.org/wiki/Nanotomography | 08:12 |
| kanzure | "The SkyScan-2011 [1] has a range of about 150 to 250 nanometers per pixel with a resolution of 400 nm and a field of view (FOV) of 200 micrometers. The Xradia nanoXCT [2] has a spatial resolution of better than 50 nm and a FOV of 16 micrometers.[1]" | 08:12 |
| kanzure | "At the Ghent University, the UGCT team developed a nano-CT scanner based on commercially available components. The UGCT facility is an open nano-CT facility giving access to scientists from universities, institutes and industry. More information can be found at UGCT-website." | 08:12 |
| fenn | also wrt scanning living brains, that's a fucking huge data rate, and at modern computing bandwidths you'd be sitting in the scanner until you die of old age | 08:13 |
| kanzure | so i think you would need to resolve individual neurons, dendrites and axons while also grabbing the synaptic weights. because if you do this in a separate pass or separate scan (especially if it was on a different day) then you will have a very big puzzle to figure out... | 08:13 |
| fenn | .wa cubic nanometers per liter | 08:13 |
| yoleaux | fenn: Sorry, no result! | 08:13 |
| fenn | let's call it around a septillion | 08:14 |
| fenn | i don't know what comes after exa- | 08:15 |
| kanzure | if that's really the bottleneck then you would have to do inline/online data compression from the results, e.g. fuzzy storage of cell membrane shape using real-time image-based segmentation of membranes from the scan data | 08:15 |
| kanzure | so.. a bunch of custom electronics. | 08:15 |
| kanzure | i don't think CT can do multiple contrast agents at the same time, right? | 08:16 |
| fenn | assuming a 1 cubic nanometer pixel size you'd need to massage around 1 million exabytes of data per channel? what's the maximum acceptable pixel size? | 08:16 |
| fenn | yottabyte | 08:17 |
| kanzure | i suppose you could do membrane labeling + one receptor in a single pass. the receptors are going to be clustered in very specific locations. and you could visually separate those from membrane. this is useful in the case of not having multiple "color" signals. | 08:17 |
| kanzure | and then if you have a different scan after N hours (to let the aptamers or whatever degrade and float out of the system), you would have to do some sort of fuzzy matching on each of the 120 different passes, while capturing all the neural membranes each time, then do some sort of fuzzy matching on each of the data sets to do a merge (you can't do a straight-up merge because the biology has changed since your last scan) | 08:18 |
| fenn | why 120? | 08:19 |
| kanzure | i don't think you are going tobe able to visually discriminate close dendrites. the cell bodies are really packed together. the membranes are sometimes literally touching. so you wont be able to get the actual connectivity from this. i just don't see how. | 08:20 |
| kanzure | he literally counted the number of known receptors and made a spreadsheet :) | 08:20 |
| fenn | but we know a lot about where cell types are located and surely don't need to do one scan per receptor type | 08:20 |
| kanzure | or at least the 120 he's interested in | 08:20 |
| kanzure | anyway, you just wont get connectivity data out of this, i don't see how to do it | 08:21 |
| fenn | you can visually discriminate dendrites if you have the cell membrane geometry and sufficient resolution | 08:21 |
| kanzure | you could do some statistical testing bullshit where you re-initialize entire simulatoins with different assumptions about connectivity until you find something that works...... but there's a lot of possibilities, and there are way too many dendrites that are very physically close to each other. (perhaps you could argue that physically close dendrites leak to each other anyway, and therefore it doesnt' matter? but this is a big assumption) | 08:21 |
| kanzure | both of the two (touching) membranes are in very close proximity | 08:22 |
| kanzure | eh i guess you could infer it..... and if you're wrong, uh.. | 08:22 |
| fenn | no i'm not saying infer it or assume they leak or anything, i'm saying visually trace them based on the geometry | 08:22 |
| fenn | geez do i need to find a picture | 08:23 |
| kanzure | your solution is basically "use even higher resolution scanning"? | 08:23 |
| kanzure | well at least we have a hypothetical alternative solution (rna/dna barcoding for connectomics) (which is lacking a speculative method for receptor density recording, although marblestone would say "just record it on dna in the cell"-- mRNA transcripts wont be enough because you need the actual density at each synapse not for the whole neuron's aggregate receptor expression with all of its umpteen thousand synapses) | 08:26 |
| fenn | i have too many tabs open apparently | 08:27 |
| kanzure | fenn: see PM | 08:27 |
| fenn | https://elife-publishing-cdn.s3.amazonaws.com/04047/elife-04047-fig5-v2.jpg (from https://elifesciences.org/content/3/e04047 ) | 08:28 |
| fenn | scale cube is 0.5 micron | 08:29 |
| fenn | 0.5 um^3 | 08:30 |
| fenn | bah who uses scale cubes | 08:30 |
| fenn | at least pick a round number like 1 | 08:30 |
| fenn | he says "synaptic weights" so i guess everyone is just crazy | 08:37 |
| kanzure | well he has also been doing some software work so i wouldn't be surprised if he says it both ways | 08:37 |
| kanzure | density/distribution is right, but the actual physical placement on the synapse is sort of irrelevant | 08:37 |
| kanzure | actually i guess we should do a simulation to confirm that irrelevance -_- | 08:38 |
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| fenn | why are they making aptamers? can't you just buy aptamers? | 08:38 |
| kanzure | aptamers for these targets do not seem to exist | 08:39 |
| kanzure | you have to make them | 08:39 |
| fenn | it's like making your own antibodies, sure you can do it but a company can do it cheaper | 08:39 |
| fenn | i guess i'm just surprised there isn't "sign here" solution for that yet | 08:40 |
| fenn | i should talk with robert mcintyre | 08:41 |
| kanzure | i wrote an intro email but haven't sent it. would you like me to send? | 08:43 |
| fenn | SBIR = small business innovation research grant | 08:44 |
| fenn | uh i am on night-time schedule currently | 08:44 |
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| kanzure | that's a stupid answer. sent. | 08:47 |
| fenn | .wik ecog | 08:53 |
| yoleaux | "Disambiguation: Ecog" — https://en.wikipedia.org/wiki/Ecog | 08:53 |
| fenn | .wik electrocorticography | 08:55 |
| yoleaux | "Electrocorticography (ECoG), or intracranial electroencephalography (iEEG), is a type of electrophysiological monitoring that uses electrodes placed directly on the exposed surface of the brain to record electrical activity from the cerebral cortex." — https://en.wikipedia.org/wiki/Electrocorticography | 08:55 |
| fenn | DNA barcoding doesn't have to be destructive, but you do have to get the DNA out of the brain cells somehow and that probably means neurons producing viral packaging proteins | 08:58 |
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| fenn | osmium | 09:01 |
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| kanzure | barcoding methods assume you have physical access to the tissue | 09:03 |
| kanzure | destructive techniques in particular | 09:03 |
| kanzure | your proposal seems to be "make sure the barcode DNA or RNA fragment gets into a virus capsid somehow"? | 09:03 |
| fenn | i'm not really sure what "barcoding" means, presumably some kind of randomly generated cell ID that you can use to reconstruct a connectome somehow | 09:04 |
| fenn | .title https://arxiv.org/abs/1404.5103 | 09:07 |
| yoleaux | [1404.5103] Rosetta Brains: A Strategy for Molecularly-Annotated Connectomics | 09:07 |
| fenn | marblestone strikes again! | 09:07 |
| kanzure | there's a recent paper too | 09:07 |
| kanzure | which cites that one | 09:07 |
| kanzure | "High-throughput mapping of single neuron projections by sequencing of barcoded RNA" http://biorxiv.org/content/early/2016/05/20/054312 | 09:07 |
| fenn | why is "in-situ sequencing" better than just random colors like brainbow? | 09:08 |
| kanzure | imaging and tracing is still a big problem | 09:08 |
| kanzure | sequencing is very cheap | 09:08 |
| fenn | uh, wtf is "in situ sequencing"? | 09:08 |
| kanzure | this changes it from an imaging and scanning problem to a sequencing problem and statistics or data mining proble | 09:08 |
| kanzure | .wik fluorescent in situ sequencing | 09:09 |
| yoleaux | "Fluorescent in situ sequencing (FISSEQ) is a method of sequencing a cells's RNA while it remains in tissue or culture using next-generation sequencing." — https://en.wikipedia.org/wiki/Fluorescent_in_situ_sequencing | 09:09 |
| kanzure | "The process is conceptually identical to the mechanism of fluorescent sequencing by synthesis in a commercial bulk DNA sequencing machine, except that it is performed in fixed tissue. Each DNA or RNA molecule in the sample is first “amplified” (i.e., copied) in-situ via rolling-circle amplification to create a localized “rolling circle colony” (rolony) consisting of identical copies of the parent molecule. A series of ... | 09:10 |
| kanzure | ... biochemical steps are then carried out. In the kth cycle, a fluorescent tag is introduced, the color of which corresponds to the identity of the kth base along the rolony’s parent DNA strand. The system is then “paused” in this state for imaging. The entire sample can be imaged in each cycle. The fluorescent tags are then cleaved and washed away, and the next cycle is initiated." | 09:10 |
| kanzure | "Each rolony – corresponding to a single “parent” DNA or RNA molecule in the tissue – thus appears across a series of fluorescent images, as a localized “spot” with a sequence of colors corresponding to the nucleotide sequence of the parent molecule. The nucleotide sequence of each DNA or RNA molecule is thus read out in-situ via fluorescent microscopy." | 09:10 |
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| fenn | ok | 09:11 |
| kanzure | there was a method like brainbow though | 09:11 |
| kanzure | "On optical detection of densely labeled synapses in neuropil and mapping connectivity with combinatorially multiplexed fluorescent synaptic markers" http://diyhpl.us/~bryan/papers2/neuro/On%20optical%20detection%20of%20densely%20labeled%20synapses%20in%20neuropil%20and%20mapping%20connectivity%20with%20combinatorially%20multiplexed%20fluorescent%20synaptic%20markers%20-%20Mishchenko%20-%202010.pdf | 09:12 |
| kanzure | i think the problem is that you run out of colors | 09:12 |
| fenn | i wonder how i had never heard of FISSEQ before | 09:14 |
| fenn | it sounds really powerful for research | 09:14 |
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| fenn | you could label each receptor with aptamers and then sequence all the aptamers in-situ to get 120 or more "colors" in your microscope image | 09:18 |
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| fenn | todd huffman's optical microtome would be enough to reconstruct a connectome that way | 09:19 |
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| kanzure | iirc you don't sequence aptamers, you just do imaging by attaching those aptamers to contrast agents | 09:23 |
| fenn | read what i said again | 09:24 |
| kanzure | oh you want to do dna hybridization imaging stuff i guess? | 09:24 |
| fenn | each aptamer is selective for a specific receptor | 09:24 |
| kanzure | yes but you don't sequence aptamers, usually | 09:24 |
| kanzure | i think there are some fluorescent hybridization techniques that you could use, where you insert your hybridization probe (another dna molecule) and then check what dna/rna/aptamer tags you have laying around in the sample | 09:25 |
| kanzure | .wik in situ hybridization | 09:25 |
| yoleaux | "In situ hybridization (ISH) is a type of hybridization that uses a labeled complementary DNA, RNA or modified nucleic acids strand (i.e., probe) to localize a specific DNA or RNA sequence in a portion or section of tissue (in situ), or, if the tissue is small enough (e.g., plant seeds, Drosophila embryos), in the entire tissue (whole …" — https://en.wikipedia.org/wiki/In_situ_hybridization | 09:25 |
| fenn | hmm the knife edge scanning microscope images a line as it moves, which wouldn't give enough time to read out an entire sequence because the position is always changes | 09:35 |
| fenn | but you could do it with a real microscope that images a whole 2d field | 09:35 |
| JayDugger | Good morning, everyone. | 09:38 |
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| kanzure | why is it a knife edge and not a laser dissector thingy? | 09:44 |
| fenn | because lasers burn stuff | 09:49 |
| fenn | i guess you could ask "why not an e-beam dissector thingy" and have a point | 09:50 |
| fenn | a knife is pretty reliable | 09:55 |
| fenn | a fast pulse of an unfocused laser would vaporize the top layer of a sample, but the material removed wouldn't be of uniform thickness because different materials (fat, water) have different heat capacities, and over time this will cause the surface to become jagged and you'd have trouble imaging it due to limited depth of field | 10:01 |
| fenn | a raster scanned focused laser would have to move very fast to prevent heat buildup in the material below what was just removed | 10:03 |
| fenn | blasting chunks of material off just seems like it would mess up the fine detail somehow | 10:04 |
| fenn | nmz787: am i mischaracterizing anything? would an ion beam or electron beam do basically the same thing? | 10:06 |
| * fenn makes an attempt to sleep | 10:14 | |
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| kanzure | "Yes, fluorescent in situ sequencing with aptamers can be done... but it's unclear what advantages over regular in situ sequencing. " | 10:29 |
| kanzure | i think the way to do it would be: dna tag <-> nanoparticle bead <-> aptamer <-> receptor where <-> is a chemical linker. | 10:29 |
| kanzure | with "regular in situ sequencing" i think you cannot tell the receptor density because the mRNA transcripts are probably not directly related to currently expressed receptos | 10:30 |
| kanzure | .. receptors. | 10:30 |
| kanzure | and the target of the in situ sequencing would be the dna fragment attached to the nanoparticle bead. (i suppose a dna fragment could be directly attached to the aptamer but i'm not familiar with the biophysics or biochemistry there and aptamer/dna interactions, since it seems like it would be dna-dna interaction) | 10:31 |
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| kanzure | fenn: one argument would be that the receptor density is so low that you would need almost single molecule PCR techniques in order to get a signal out of that mess | 10:36 |
| kanzure | bitcoin core meeting in 1 hour | 10:42 |
| kanzure | "Fluorescent in situ sequencing (FISSEQ) of RNA for gene expression profiling in intact cells and tissues" http://www.nature.com/nprot/journal/v10/n3/full/nprot.2014.191.html | 10:43 |
| kanzure | but again, i don't think gene expression profiling is the same thing as looking at the actual receptors | 10:43 |
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| kanzure | "In order to get the protein to the synapse, the receptor must be translated from the RNA. So a method of using FISSEQ for synaptic weights would be cool." | 10:45 |
| kanzure | yeah but.... i don't think that all the RNA is floating around constantly. the replacement rate for receptors might be days or weeks, not minutes. | 10:45 |
| fenn | the aptamer is a piece of DNA that is bound to a receptor, and you sequence it because it's made of DNA | 10:55 |
| fenn | no beads needed | 10:55 |
| maaku | someone needs to setup a robot with an iphone and internet connection, and do “twitch plays pokemon now" | 10:58 |
| kanzure | i don't think you can sequence aptamers directly | 11:01 |
| kanzure | you can degrade aptamers and then sequence them | 11:01 |
| kanzure | but they are supposed to be folded up | 11:02 |
| kanzure | maaku: so i was talking with the twitchplayspokemon person last night about that, he says he doesn't have enough bandwidth to pursue that :( | 11:02 |
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| kanzure | maaku: they are spending all of their cycles on finishing "pokemon prism", which they recently migrated from pokemon gold to pokecrystal source code | 11:03 |
| kanzure | maaku: my proposal to them was to have the chat/irc interface let users type in coordinates or directions, and then the video feed would have an android-x86 machine running pokemon go. and then moving left/right etc would move on google maps or google streetview. | 11:05 |
| kanzure | "Flare: An approach to routing in lightning network" http://bitfury.com/content/5-white-papers-research/whitepaper_flare_an_approach_to_routing_in_lightning_network_7_7_2016.pdf | 11:11 |
| fenn | if you can't sequence the folded up part of the aptamer then you just stick a unique tail on it | 11:13 |
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| kanzure | yes but i don't know about dna/aptamer biophysics stuff, so i just assumed you would need a nanoparticle between the aptamer and dna tag | 11:15 |
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| kanzure | "The industrial melanism mutation in British peppered moths is a transposable element" http://www.nature.com/nature/journal/v534/n7605/full/nature17951.html | 11:21 |
| kanzure | "Single-vesicle imaging reveals different transport mechanisms between glutamatergic and GABAergic vesicles" http://science.sciencemag.org/content/351/6276/981 | 11:21 |
| kanzure | just closing out some tabs :\ | 11:21 |
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| kanzure | https://www.technologyreview.com/s/532166/with-100-million-entrepreneur-sees-path-to-disrupt-medical-imaging/ | 13:58 |
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| kanzure | https://www.butterflynetinc.com/#What | 14:02 |
| kanzure | "At Butterfly Network, we are reinventing the ultrasound machine by squeezing all of its components onto a single silicon chip. " | 14:02 |
| kanzure | wasn't this the same guy from 454 that was spending a lot of money on finding genes related to 'mathematical genius' phenotypes | 14:02 |
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| kanzure | (jonathan rothberg) | 14:03 |
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| kanzure | .wik capacitive micro-machined ultrasound transducers | 14:29 |
| yoleaux | kanzure: Sorry, that command (.wik) crashed. | 14:29 |
| kanzure | .wik capacitive micro-machined ultrasound transducers | 14:29 |
| yoleaux | kanzure: Sorry, that command (.wik) crashed. | 14:29 |
| kanzure | https://en.wikipedia.org/wiki/Capacitive_micromachined_ultrasonic_transducers | 14:30 |
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| kanzure | .title https://www.youtube.com/watch?v=2yfXgu37iyI | 15:11 |
| yoleaux | Doctor Strangelove - Doomsday Machine - YouTube | 15:11 |
| chris_99 | seen this kanzure https://en.wikipedia.org/wiki/Dead_Hand_%28nuclear_war%29 | 15:12 |
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| kanzure | .title https://www.youtube.com/watch?v=BArDvf9_U90 | 15:30 |
| yoleaux | IndieBio SF Live Stream Demo Day 3 (7/14/2016) - YouTube | 15:30 |
| kanzure | .title https://www.youtube.com/watch?v=YJDoGtHpn5I | 15:31 |
| yoleaux | IndieBio SF Live Stream Demo Day 3 (7/14/2016) - YouTube | 15:31 |
| kanzure | mycoworks doing some leather stuff | 15:32 |
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| nmz787_i | .wik cryoelectrontomography | 17:02 |
| yoleaux | nmz787_i: Sorry, that command (.wik) crashed. | 17:02 |
| nmz787_i | .wik cryo-electron tomography | 17:02 |
| yoleaux | "Electron cryo-tomography (ECT, also called cryo-electron tomography, cryo-ET or CET) is an imaging technique used to produce high-resolution (~4 nm) three-dimensional views of samples, typically biological macromolecules and cells." — https://en.wikipedia.org/wiki/Cryo-electron_tomography | 17:02 |
| nmz787_i | you might die waiting, but as long as the brain was kept in good cryo... the results wouldn't be messed up | 17:03 |
| kanzure | nmz787_i: fenn asked earlier today about using e-beams for dissecting brain tissue. does it burn or is it a clean cut? | 17:03 |
| nmz787_i | start now for your grandkids! | 17:03 |
| nmz787_i | hrmm, I am not sure what an e-beam would do... I assume it would be very slow... it is going to definitely cause bonds to break and maybe some to form... so I guess that is 'burning' though I am not sure how much offgassing would occur... I imagine it wouldn't be too different than 'normal burning' | 17:05 |
| nmz787_i | .wik Serial block-face scanning electron microscopy | 17:05 |
| yoleaux | "Serial block-face scanning electron microscopy (SBEM, SBSEM or SBFSEM) is a method to generate high resolution three-dimensional images from small samples, often biological samples such as brain tissue. A serial block-face scanning electron microscope consists of an ultramicrotome mounted inside the vacuum chamber of a scanning electron …" — https://en.wikipedia.org/wiki/Serial_block-face_scanning_electron_microscopy | 17:05 |
| nmz787_i | prep for either of those techniques is often a FIB | 17:05 |
| kanzure | specifically the point is that if you break too much of the neural tissue then you can't reconnect the neurals after you are done imaging | 17:06 |
| nmz787_i | and in the latter, a dual-beam SEM+FIB is used, FIB for fine-slicing, SEM for imaging | 17:06 |
| nmz787_i | ah | 17:06 |
| kanzure | okay fine. FIB then. but why is todd/3scan using a knife if FIB is better ? | 17:06 |
| nmz787_i | well that is mostly automated with the latter wiki tech | 17:06 |
| nmz787_i | knife is cheaper and faster | 17:06 |
| nmz787_i | maybe he wants/needs less resolution | 17:06 |
| kanzure | cost of FIB is the machine itself ya? | 17:06 |
| nmz787_i | hmm, I guess that is definitely part of it | 17:07 |
| nmz787_i | consumables are expensive, but you would take a while to consume the consumables (i.e. a year probably of heavy use) | 17:07 |
| nmz787_i | (i.e. a new Gallium FIB tip) | 17:07 |
| nmz787_i | or a new aperture strip (cuts off the main beam, to make the spot smaller and finer.... i.e. go from a fat-marker to a crayon to a fine-tip-pencil) | 17:09 |
| kanzure | btw don't worry about spilling the beans on your idea. the guy wasn't even listening :\. | 17:10 |
| nmz787_i | ok | 17:10 |
| nmz787_i | also you can probably distinguish some receptor or membrane differences with either x-ray spectroscopy (EDS or EDX) or maybe EELS (electron energy loss spectroscopy) | 17:11 |
| nmz787_i | .wik microprobe | 17:11 |
| yoleaux | "A microprobe is an instrument that applies a stable and well-focused beam of charged particles (electrons or ions) to a sample." — https://en.wikipedia.org/wiki/Microprobe | 17:11 |
| nmz787_i | .wik edx | 17:12 |
| yoleaux | "EDX may refer to:" — https://en.wikipedia.org/wiki/Edx | 17:12 |
| nmz787_i | heh | 17:12 |
| nmz787_i | .wik eels | 17:12 |
| yoleaux | "An eel is any fish belonging to the order Anguilliformes (/æŋˌɡwɪlᵻˈfɔːrmiːz/), which consists of four suborders, 20 families, 111 genera and about 800 species. Most eels are predators." — https://en.wikipedia.org/wiki/Eels | 17:12 |
| nmz787_i | haha | 17:12 |
| nmz787_i | .wik electron energy loss spectroscopy | 17:12 |
| yoleaux | "In electron energy loss spectroscopy (EELS) a material is exposed to a beam of electrons with a known, narrow range of kinetic energies. Some of the electrons will undergo inelastic scattering, which means that they lose energy and have their paths slightly and randomly deflected." — https://en.wikipedia.org/wiki/Electron_energy_loss_spectroscopy | 17:12 |
| nmz787_i | .wik energy dispersive x-ray | 17:12 |
| yoleaux | nmz787_i: Sorry, that command (.wik) crashed. | 17:12 |
| nmz787_i | .wik Energy-dispersive X-ray spectroscopy | 17:12 |
| yoleaux | "Energy-dispersive X-ray spectroscopy (EDS, EDX, or XEDS), sometimes called energy dispersive X-ray analysis (EDXA) or energy dispersive X-ray microanalysis (EDXMA), is an analytical technique used for the elemental analysis or chemical characterization of a sample. It relies on an interaction of some source of X-ray excitation and a …" — https://en.wikipedia.org/wiki/Energy-dispersive_X-ray_spectroscopy | 17:12 |
| nmz787_i | http://www.wormatlas.org/EMmethods/FIBSEM.htm | 17:15 |
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| kanzure | "Sequencing the connectome" (2012) http://journals.plos.org/plosbiology/article?id=10.1371/journal.pbio.1001411 | 17:43 |
| kanzure | "Connectivity determines the function of neural circuits. Historically, circuit mapping has usually been viewed as a problem of microscopy, but no current method can achieve high-throughput mapping of entire circuits with single neuron precision. Here we describe a novel approach to determining connectivity. We propose BOINC (“barcoding of individual neuronal connections”), a method for converting the problem of connectivity into a ... | 17:43 |
| kanzure | ... form that can be read out by high-throughput DNA sequencing. The appeal of using sequencing is that its scale—sequencing billions of nucleotides per day is now routine—is a natural match to the complexity of neural circuits. An inexpensive high-throughput technique for establishing circuit connectivity at single neuron resolution could transform neuroscience research." | 17:43 |
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| nmz787 | I remember that BOINC acronym | 17:58 |
| nmz787 | I seem to recall that was a clever paper | 17:59 |
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| streety | it's also the software behind seti@home, rosetta@home, etc | 18:09 |
| kanzure | yes that's something else. | 18:12 |
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| nmz787 | hmm, India has 4x the population, but 7x the car accident death rate | 18:15 |
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| kanzure | w | 19:30 |
| kanzure | hmph | 19:30 |
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| kanzure | "In vivo generation of DNA sequence diversity for cellular barcoding" http://zadorlab.cshl.edu/PDF/peikon-zador2014.pdf (2014) | 23:06 |
| kanzure | "We have developed a novel method of uniquely tagging individual cells in vivo with a genetic ‘barcode’ that can be recovered by DNA sequencing. Our method is a two-component system comprised of a genetic barcode cassette whose fragments are shuffled by Rci, a site-specific DNA invertase. The system is highly scalable, with the potential to generate theoretical diversities in the billions. We demonstrate the feasibility of this ... | 23:07 |
| kanzure | ... technique in Escherichia coli. Currently, this method could be employed to track the dynamics of populations of microbes through various bottlenecks." | 23:07 |
| kanzure | "Puzzle imaging: using large-scale dimensionality reduction algorithms for localization" http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0131593 | 23:09 |
| kanzure | "We here present a novel approach, “puzzle imaging,” that allows imaging a spatially scrambled sample. This technique takes many spatially disordered samples, and then pieces them back together using local properties embedded within the sample. We show that puzzle imaging can efficiently produce high-resolution images using dimensionality reduction algorithms. We demonstrate the theoretical capabilities of puzzle imaging in three ... | 23:09 |
| kanzure | ... biological scenarios, showing that (1) relatively precise 3-dimensional brain imaging is possible; (2) the physical structure of a neural network can often be recovered based only on the neural connectivity matrix; and (3) a chemical map could be reproduced using bacteria with chemosensitive DNA and conjugative transfer. The ability to reconstruct scrambled images promises to enable imaging based on DNA sequencing of homogenized ... | 23:09 |
| kanzure | ... tissue samples." | 23:09 |
| kanzure | "spatially scrambled sample" like eggs | 23:09 |
| kanzure | "Site-specific recombinatorics: in situ cellular barcoding with the Cre Lox system" http://arxiv.org/pdf/1602.02190.pdf (2016) | 23:11 |
| kanzure | "Cellular barcoding is a significant, recently developed, biotechnology tool that enables the familial identification of progeny of individual cells in vivo. Most existing approaches rely on ex vivo viral transduction of cells with barcodes, followed by adoptive transfer into an animal, which works well for some systems, but precludes barcoding cells in their native environment, such as those inside solid tissues. With a view to ... | 23:11 |
| kanzure | ... overcoming this limitation, we propose a new design for a genetic barcoding construct based on the Cre Lox system that induces randomly created stable barcodes in cells in situ by exploiting inherent sequence distance constraints during site-specific recombination. Leveraging this previously unused feature, we identify the cassette with maximal code diversity. This proves to be orders of magnitude higher than what is attainable with ... | 23:11 |
| kanzure | ... previously considered Cre Lox barcoding approaches and is well suited for its intended applications as it exceeds the number of lymphocytes or hematopoietic progenitor cells in mice. Moreover, it can be built using established technology." | 23:11 |
| kanzure | "Internal readout system for molecular recorders" http://ieeexplore.ieee.org/xpls/abs_all.jsp?arnumber=7181696 (2015, Magierowski and Messier) | 23:12 |
| kanzure | "To simplify the problem this work discusses a centralized means of achieving readout that leverages an organism's circulatory system and sensors for DNA sequencing. It studies a scenario where molecular recordings from the brain are allowed to circulate in the bloodstream while nanopores arrayed in vessels near the heart pick-up and process the messages. The circulatory system serves as a means of transport and signal redundancy while ... | 23:13 |
| kanzure | ... the sequencing technique offers the possibility of a monolithic high-throughput single-molecule detector." | 23:13 |
| kanzure | .wik Hin recombinase | 23:15 |
| yoleaux | "Hin recombinase is a 21kD protein composed of 198 amino acids that is found in the bacteria Salmonella. Hin belongs to the serine recombinase family of DNA invertases in which it relies on the active site serine to initiate DNA cleavage and recombination." — https://en.wikipedia.org/wiki/Hin_recombinase | 23:15 |
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| kanzure | will be trivial to deliver new barcoding systems to all the neurons with viruses or something. obvious thing to deliver would be optimized recombinases etc. | 23:16 |
| kanzure | if you could somehow copy a barcode to each outgoing message then you have a cellular debugging system ready to go... but i don't know how to correctly attach a copy of a cell's dna or rna barcode molecule. i guess you could use a hacky crispr system where you try to splice in a matching fragment, into the outgoing message. and then some messages wont have the additional address/barcode data but that's kinda okay. | 23:19 |
| kanzure | specifically, the barcode would have to be specified in a dna molecule in such a way that when the crispr system hack is expressed, it naturally knows the barcode and to put that in each of the outgoing messages. | 23:20 |
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| kanzure | "A new defective helper RNA to produce recombinant Sindbis virus that infects neurons but does not propagate" http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4877524/ | 23:25 |
| kanzure | "Long-term Cre-mediated retrograde tagging of neurons using a novel recombinant pseudorabies virus" http://journal.frontiersin.org/article/10.3389/fnana.2014.00086/full | 23:27 |
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| kanzure | "A single adaptive point mutation in Japanese encephalitis virus capsid is sufficient to render the virus as a stable vector for gene delivery" | 23:28 |
| kanzure | "Large-scale fluorescence calcium-imaging methods for studies of long-term memory in behaving mammals" http://pyramidal.stanford.edu/publications/Jercog2016-CSHPB.pdf (2016) | 23:29 |
| kanzure | page 10 figure 3 shows their setup for fluoresence microscopy in mice | 23:29 |
| kanzure | "Neuroanatomy goes viral" http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4486834/ (2015) | 23:31 |
| kanzure | "Fluorescent in situ sequencing (FISSEQ) of RNA for gene expression profiling in intact cells and tissues" http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4327781/ (2016, church etc) | 23:37 |
| kanzure | hah! | 23:39 |
| kanzure | greg fahy interviews george church http://www.lifeextension.com/Lpages/2016/CRISPR/index | 23:39 |
| kanzure | "Androcyte has also received two elderly Arabian mares 28 and 30 years old (age-equivalent to 80-year-old humans) from a sanctuary. If genes delivered by CRISPR to the mice are able to restore youth and health, CRISPR delivery of those genes will be tested on the horses to show that large animals can also benefit." | 23:41 |
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| kanzure | also i know we have already been over this but it's still cool "Saturated reconstruction of a volume of neocortex" https://www.mcb.harvard.edu/mcb_files/media/editor_uploads/2015/07/PIIS0092867415008247.pdf | 23:43 |
| kanzure | https://scholar.google.com/scholar?cites=9974693126559535546&as_sdt=5,44&sciodt=0,44&hl=en | 23:44 |
| kanzure | "Nanoconnectomic upper bound on the variability of synaptic plasticity" https://elifesciences.org/content/4/e10778?dom=pscau&src=syn | 23:45 |
| kanzure | "We found that there is a minimum of 26 distinguishable synaptic strengths, corresponding to storing 4.7 bits of information at each synapse." | 23:45 |
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| kanzure | i still don't have a copy of "Simple, Scalable Proteomic Imaging for High-Dimensional Profiling of Intact Systems" http://www.sciencedirect.com/science/article/pii/S0092867415015056 | 23:48 |
| kanzure | "With SWITCH, we performed combinatorial protein expression profiling of the human cortex and also interrogated the geometric structure of the fiber pathways in mouse brains." | 23:49 |
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| kanzure | fenn: ^that seems to be the same proposal. | 23:49 |
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