| kanzure | hi all | 01:56 |
|---|---|---|
| kristianpaul | hey | 02:22 |
| kanzure | hi azonenberg | 04:27 |
| gene_hacker | anyone home? | 04:29 |
| gene_hacker | wondering how you guys make your masks | 04:30 |
| gene_hacker | or are you using a DLP projector to do maskless lithography? | 04:31 |
| azonenberg | hi kanzure | 06:07 |
| azonenberg | whoops, seems i missed somebody | 06:07 |
| azonenberg | Was having fun with BGA soldering in the living room | 06:07 |
| azonenberg | and in case he comes back, i use a normal office laser printer | 06:07 |
| azonenberg | then reduce 10x with a microscope objective | 06:07 |
| kanzure | azonenberg: http://www.digikey.com/product-highlights/us/en/texas-instruments-pico/688 | 06:46 |
| kanzure | i'm thinking about getting a dmd dev kit | 06:46 |
| kanzure | it would be especially helpful to have find one that does UV | 06:47 |
| azonenberg | kanzure: link doesnt work for me | 06:47 |
| azonenberg | its a 404 | 06:47 |
| kanzure | works for me o.o | 06:47 |
| kanzure | maybe you block digikey. i wouldn't blame you. | 06:47 |
| azonenberg | lol | 06:47 |
| azonenberg | i see a dk 404 page | 06:47 |
| azonenberg | i buy stuff from them all the time | 06:47 |
| kanzure | http://www.ti.com/tool/dlpd4x00kit | 06:48 |
| kanzure | are you doing maskless lithography? | 06:48 |
| azonenberg | No | 06:48 |
| kanzure | fuck | 06:48 |
| azonenberg | Print on laser printer transparency | 06:48 |
| azonenberg | reduce 10x optically | 06:48 |
| azonenberg | i am looking at doing laser direct write with a bluray diode | 06:48 |
| azonenberg | straight from the GDS | 06:48 |
| azonenberg | But thats a ways out | 06:48 |
| kanzure | alright | 06:48 |
| kanzure | i'm doing a dna synthesis chip | 06:49 |
| azonenberg | Ooh | 06:49 |
| kanzure | http://diyhpl.us/~bryan/papers2/microfluidics/ | 06:49 |
| kanzure | http://diyhpl.us/~bryan/papers2/DNA/ | 06:49 |
| kanzure | http://diyhpl.us/~bryan/papers2/microfluidics/synthesis/ | 06:49 |
| kanzure | also, any papers you can contribute on #homecmos would be welcomed | 06:49 |
| azonenberg | very nice | 06:50 |
| azonenberg | and well, my work has actually mostly been in MEMS | 06:50 |
| azonenberg | my lab notes are in the google code repository in the topic | 06:50 |
| kanzure | etching processes are pretty similar in a few places | 06:50 |
| kanzure | sometimes microfluidics requires gold electrodes and other crap | 06:51 |
| azonenberg | Yeah | 06:51 |
| kanzure | but has the same geometry/layout problems that you need to consider | 06:51 |
| azonenberg | And larger | 06:51 |
| azonenberg | Which is a huge benefit for home / low budget fab | 06:51 |
| kanzure | yes, well, i'm not complaining about that | 06:51 |
| comeon | why are nitrides like silicon nitride or titanium nitride such good "barrier" layer materials? | 07:43 |
| azonenberg | comeon: Mostly because they're denser, i think | 07:45 |
| azonenberg | I'm a little less up on the transistor side of things as i've mostly studied MEMS | 07:45 |
| azonenberg | I dont see Si3N4 used as a barrier as much | 07:45 |
| azonenberg | TiN and TaN i see used | 07:45 |
| comeon | nonstoichiometric silicon nitride is pecvd at low temperatures on gaas after doping gaas so that during an lattice damage removal annealing heat treatment the as doesn't float away | 07:47 |
| comeon | fuckin nazis on ##chemistry and ##physics quieted me | 07:48 |
| azonenberg | I assume by non-stoichiometric they mean Si rich and not N rich | 07:48 |
| azonenberg | but i'm not sure of the exact characteristics of pecvd nitride's formulation | 07:48 |
| azonenberg | but makes sense | 07:48 |
| azonenberg | also just fyi this is the unofficial microfab / semiconductor channel on freenode | 07:50 |
| azonenberg | not just home fab | 07:50 |
| comeon | then rename it | 07:50 |
| azonenberg | I might | 07:50 |
| azonenberg | Just saying, discussion of cleanroom fab on "real" tools is not offtopic | 07:50 |
| comeon | well i only asked because the bigger channels quieted me | 07:51 |
| comeon | but you're the only person who ever talks here anyway | 07:51 |
| comeon | might as well call it #azonenberg | 07:51 |
| azonenberg | Lol | 07:52 |
| azonenberg | Other people do talk | 07:52 |
| azonenberg | But they idle a lot | 07:52 |
| azonenberg | the community is small | 07:52 |
| azonenberg | Not many people interested in semiconductors | 07:53 |
| azonenberg | and even less know anything about them | 07:53 |
| azonenberg | even less are on freenode | 07:53 |
| comeon | freenode has a lot of software cocksuckers | 07:53 |
| azonenberg | lol | 07:53 |
| azonenberg | Hey, dont knock software TOO much | 07:53 |
| azonenberg | my undergraduate degree is in computer science | 07:54 |
| comeon | psh | 07:54 |
| azonenberg | and i'm going for a PhD in the same | 07:54 |
| comeon | why | 07:54 |
| azonenberg | My research is in computer architecture | 07:54 |
| azonenberg | and operating system security | 07:54 |
| comeon | that sounds super boring | 07:54 |
| azonenberg | Not at all | 07:54 |
| azonenberg | my advisor is a crypto guy and i'm not too into the intense theory but when it comes time to prototype stuff its fun | 07:54 |
| comeon | whatever floats your boat dude | 07:55 |
| azonenberg | I just do hardware stuff for fun lol | 07:55 |
| comeon | there is a dude in ##electronics who literally floats in a boat | 07:55 |
| azonenberg | starting to take EE classes because i've taken most of the ones CS offers adn am not interested in the rest | 07:55 |
| comeon | geckosenator or something | 07:55 |
| azonenberg | interesting, i've heard of the guy but didnt know he lived on a boat | 07:56 |
| comeon | he cooks his own shit for fuel | 07:56 |
| azonenberg | lol | 07:56 |
| comeon | super greenie weirdo | 07:56 |
| soul-d | lol | 07:56 |
| soul-d | i din't want to know that | 07:56 |
| azonenberg | i assume he's online via a solar-powered laptop | 07:56 |
| azonenberg | with a satellite or 3g connection? | 07:56 |
| comeon | and the asshole lives on food stamps | 07:56 |
| comeon | cuz he doesn't want to work a real job | 07:57 |
| comeon | just float on a boat and leech off society | 07:57 |
| azonenberg | furrywolf is mostly off-grid too, he/she (i think wolfy is a female) runs almost entirely on solar | 07:57 |
| soul-d | i think most people in here are just to steal azonenberg's idea's for world domination | 07:57 |
| soul-d | at least i am | 07:57 |
| azonenberg | soul-d: they'll be waiting a long time | 07:57 |
| comeon | got to milk them nerds | 07:57 |
| soul-d | no key is nano bots :P | 07:57 |
| azonenberg | this isnt ##world-domination | 07:57 |
| azonenberg | my world domination plans are occurring elsewhere | 07:57 |
| comeon | i'm gonna get shiftaced | 07:57 |
| comeon | tired as fuck already | 07:57 |
| azonenberg | comeon: what real nerds do with alcohol https://fbcdn-sphotos-a.akamaihd.net/hphotos-ak-snc7/292037_2309167690240_1280772249_2764502_5609467_n.jpg | 07:58 |
| azonenberg | (me on my 21st birthday) | 07:58 |
| comeon | seriously? | 07:58 |
| azonenberg | Yes | 07:58 |
| comeon | you need a life son | 07:59 |
| soul-d | lol he got one | 07:59 |
| azonenberg | i dont drink but i wanted to get a picture of me and a bottle of alcohol on my 21st | 07:59 |
| soul-d | he does what he likes | 07:59 |
| comeon | you don't drink? | 07:59 |
| comeon | like at all? | 07:59 |
| azonenberg | Nope | 07:59 |
| comeon | why not? | 07:59 |
| azonenberg | totally clean | 07:59 |
| comeon | are you a moralfag? | 07:59 |
| azonenberg | and i just dont feel the need to alter my neurotransmitters at all | 07:59 |
| azonenberg | i like 'em just the way they are | 07:59 |
| azonenberg | if it's not broken dont fix it | 07:59 |
| comeon | but it's fun | 08:00 |
| azonenberg | anyway so i showed the pic to my doctoral advisor and he thought it was hilarious | 08:00 |
| azonenberg | comeon: maybe you think it is, and if so you're welcome to do it | 08:00 |
| azonenberg | while you're off puking your guts up i'll be doing FPGA design | 08:00 |
| comeon | have you tried it at least? | 08:00 |
| comeon | you don't have to drink to you puke your guts | 08:00 |
| azonenberg | I know i wouldnt like it | 08:01 |
| azonenberg | my mind cant handle downtime | 08:01 |
| azonenberg | after i graduated college i went with my family down the shore for a vacation | 08:01 |
| azonenberg | By the end of the second or third day i couldnt take it anymore | 08:01 |
| azonenberg | pulled out my laptop and started doing mask layout for a MEMS comb drive | 08:01 |
| comeon | you need to get laid | 08:01 |
| azonenberg | i need to be doing something intellectual every waking hour, basically, or i go nuts | 08:01 |
| kanzure | i don't talk much in here because of the other diybio channel ##hplusroadmap | 08:02 |
| kanzure | but that one is called ##kanzure not #azonenberg | 08:02 |
| azonenberg | lol | 08:02 |
| kanzure | also, that channel is full of neurofags doing nootropics | 08:02 |
| kanzure | but i think you were talking about alcohol | 08:03 |
| kanzure | so nevermind | 08:03 |
| Action: kanzure sleeps | 08:03 | |
| comeon | what | 08:03 |
| comeon | i don't know man. you need to lighten up | 08:03 |
| azonenberg | comeon: lighten up? What is that going to do for me | 08:04 |
| azonenberg | i'm 21, most of my friends havent even graduated yet, and i have companies beating a path to my door basically offering me jobs | 08:04 |
| comeon | there's more to life than companies and stuff | 08:05 |
| azonenberg | i'm in a PhD program at a well known engineering school doing work i love, with enough spare time to do crazy research and projects on the side | 08:05 |
| azonenberg | comeon: suffice it to say, ten years from now i think i'll be very happy with where i am | 08:08 |
| comeon | where is that? | 08:08 |
| soul-d | in a position he can look back | 08:08 |
| soul-d | and be proud on | 08:09 |
| soul-d | instead of wasted time | 08:09 |
| soul-d | drinking | 08:09 |
| soul-d | complaining | 08:09 |
| soul-d | telling others tehy don't have a lvie becasue you don;t have one | 08:09 |
| azonenberg | High paying job at a R&D lab for some big tech company or national lab, living in a house/lab i built from the ground up with my own two hands | 08:09 |
| azonenberg | loaded with tools and equipment rivaling a small university | 08:09 |
| azonenberg | all the toys i could ever want, from a SEM to a class 1000 cleanroom to a machine shop | 08:09 |
| azonenberg | by that point i will likely have large submicron (750nm or so) CMOS fab working | 08:10 |
| azonenberg | oh, and out in the country so i can enjoy the outdoors and not be bothered by neighbors living too close | 08:11 |
| soul-d | azonenberg, whenever your company put's out stocks give me pm id like to take some | 08:11 |
| azonenberg | though still within a commuting distance of wherever i work | 08:11 |
| azonenberg | soul-d: lol, the last company i owned closed its doors when my partner took a full time offer and i left for grad school | 08:11 |
| azonenberg | so thats a ways out | 08:11 |
| comeon | do you have friends in real life? | 08:11 |
| azonenberg | comeon: sure - my roommates, most of the electronics club | 08:12 |
| soul-d | given your personality comeon i doubt you have any real friends | 08:12 |
| azonenberg | i dont go out drinking with them, more likely i'll wander into the living room at 2 AM and find one of them working in the lab | 08:12 |
| soul-d | just friendly to your face till you are gone | 08:12 |
| azonenberg | or etching a PCB in the fume hood | 08:12 |
| azonenberg | And you know what? I wouldnt have it any other way | 08:13 |
| comeon | soul-d: i don't get teetotalers | 08:15 |
| soul-d | who said he was and why are you obseesed by it | 08:16 |
| soul-d | maybe he understands the chemical formula | 08:16 |
| soul-d | and knows it's basicaly poision | 08:16 |
| azonenberg | Also, for the record, i made this FPGA dev board http://i.imgur.com/6pYLF.jpg today to relax | 08:17 |
| comeon | in small quantities it's healthier than completely abstaining | 08:17 |
| azonenberg | and get away from studying for my upcoming PhD qualifying exam | 08:17 |
| soul-d | did he say he did and why are you intrested in why are you obsessed with an online persons life ? | 08:18 |
| soul-d | and why do you want to change it | 08:18 |
| soul-d | somthing to do with just respecting people's choices | 08:18 |
| comeon | what's with the whiteknight? | 08:19 |
| azonenberg | i dont need anyone to defend me, he's just having fun poking the troll | 08:20 |
| comeon | you don't have to respect people's choices especially if they are super different than everyone else | 08:20 |
| Action: azonenberg pops lid of "troll food" can and throws it toward ##conspiracy-theory | 08:21 | |
| soul-d | i don't take people that use the word "super" very seriously | 08:21 |
| comeon | i don't take people that use the word "very" super seriously | 08:21 |
| azonenberg | And for the record i do not care what you think about me whatsoever, i am perfectly happy with where i am in life and am not "missing out" on anything | 08:22 |
| azonenberg | as a result of spending my evenings designing microprocessors instead of going out drinking | 08:22 |
| soul-d | one establisches brain networks and makes you measurably smarter other | 08:23 |
| soul-d | ... | 08:23 |
| soul-d | you can guess | 08:23 |
| azonenberg | If you enjoy going out and ingesting near-toxic levels of ethanol, thats your decision | 08:24 |
| azonenberg | One less person to compete for my job | 08:24 |
| azonenberg | But i find joy in intellectual stimulation and there's absolutely nothing wrong with that | 08:25 |
| soul-d | and i can tell from experience | 08:27 |
| soul-d | that it's better tehn waking up in a ditch | 08:27 |
| soul-d | afhter to much alchol | 08:27 |
| soul-d | still wonder how i got home | 08:28 |
| soul-d | alive | 08:28 |
| soul-d | i hate missing parts of memory :P | 08:28 |
| lekernel_ | ahem | 08:51 |
| lekernel_ | http://thebestpageintheuniverse.net/c.cgi?u=math | 08:51 |
| lekernel_ | http://www.paulgraham.com/icad.html | 08:51 |
| Sync | kanzure: I probably can get you a DLP devkit | 11:25 |
| kanzure | Sync: what are the details | 18:22 |
| kanzure | also, can you possibly get me one that can also do UV | 18:23 |
| Sync | they are sillicon imaging dev boards iirc | 18:23 |
| kanzure | azonenberg: i'm a little disappointed that you're in school :) | 18:24 |
| azonenberg | kanzure: why? Lol | 18:24 |
| azonenberg | thought i was some 14-year-old kid? | 18:24 |
| kanzure | Sync: link? | 18:24 |
| kanzure | azonenberg: that would be the ideal case yes | 18:25 |
| azonenberg | Lol | 18:25 |
| kanzure | azonenberg: but more to the point that you feel university is necessary at all | 18:25 |
| azonenberg | Oh | 18:25 |
| azonenberg | I needed a piece of paper that says "bachelor of science" to get some employers to even look at me | 18:25 |
| azonenberg | cant say i've learned much from class, i dont find lectures a good way to learn | 18:25 |
| kanzure | are those the types of people that you really want to work for | 18:26 |
| kanzure | your employment requirements should just be money, imho | 18:26 |
| azonenberg | No, but | 18:26 |
| azonenberg | they'd pay the bills while i looked for the *right* people | 18:26 |
| Sync | I don't know which ones gotta find them first | 18:27 |
| kanzure | i saw some pretty cheap dlp dev kits for $300ish | 18:27 |
| kanzure | but they didn't do UV | 18:27 |
| kanzure | UV would be nice so that i can do DNA quantification by raman absorption spectroscopy or something | 18:28 |
| azonenberg | They specifically said no uv, or they didnt say they did? | 18:28 |
| azonenberg | And depends on the wavelength | 18:28 |
| kanzure | their support varies, i've looked at some spec sheets.. whatever | 18:28 |
| azonenberg | for lithography using DNQ-novolac photoresist you can get stuff that responds at 405 or even 450nm (my resist is 405 peak) | 18:28 |
| kanzure | there's a few problems with dna synthesis, maybe you have some ideas | 18:28 |
| azonenberg | which is actually on the edge of the visible spectrum | 18:29 |
| kanzure | if i explain what the process is. | 18:29 |
| azonenberg | Well, i understand at a very basic level how dna replication works | 18:29 |
| azonenberg | but when you say synthesis | 18:29 |
| kanzure | nah this is different | 18:29 |
| kanzure | *different | 18:29 |
| azonenberg | you want to take a bunch of raw nucleotides and make a piece of dna? | 18:29 |
| azonenberg | with a specific sequence? | 18:29 |
| kanzure | you can use dna polymerase to make new sequences but you can't control what nucleotides it chooses | 18:29 |
| kanzure | there's a few different chemistries for adding nucleotides to a growing single strand | 18:29 |
| kanzure | this is generally called oligonucleotide synthesis | 18:29 |
| kanzure | the most popular solid phase oligonucleotide synthesis method is phosphoramidite chemistry | 18:30 |
| azonenberg | ok | 18:30 |
| kanzure | there's also a version of this phosphoramidite chemistry that uses photolabile protecting groups on the ddNTPs | 18:30 |
| azonenberg | Bear in mind my ochem knowledge is relatively limited | 18:30 |
| azonenberg | i have more experience with inorganic and even that is somewhat limitd | 18:30 |
| kanzure | so you shine some light to decap, and hope a reaction occurs to add that nucleotide to the end of the growing strand | 18:31 |
| kanzure | wash step. then some other capping/decapping steps. | 18:31 |
| kanzure | the wash step is the main pain in the ass for me | 18:31 |
| kanzure | it's not a complicated step, but i mean from a practical standpoint | 18:31 |
| azonenberg | Ok, let me see if i have this straight | 18:32 |
| azonenberg | Start with a single nucleotide, attach chemical A to one end and B to the other | 18:32 |
| azonenberg | B is stable and A degrades under UV | 18:32 |
| azonenberg | Use UV to remove A, add a small amount of your second nucleotide and attach to that end of the strand | 18:33 |
| azonenberg | remove unreacted nucleotide somehow, add more A to cap it? | 18:33 |
| kanzure | more or less, yes | 18:33 |
| kanzure | "remove unreacted nucleotide" is a giant wash step | 18:34 |
| azonenberg | Yeah, i can imagine that'd be fun to do without taking out your target strands | 18:34 |
| azonenberg | no clue where to start myself lol | 18:34 |
| kanzure | most microfluidic devices use continuous flow, so they just flow the wash chemistry by all of the 'array units' | 18:34 |
| azonenberg | Anyway so why do you need lithography-style patterning for this? | 18:34 |
| azonenberg | are you talking for building a microfluidic unit? | 18:34 |
| kanzure | most of the time this chemistry is done on solid supports (beads) | 18:34 |
| kanzure | so you can imagine an array of beads and DLP/DMD using different wavelengths to activate different nucleotides at different locations (beads in physically separate chambers) | 18:35 |
| kanzure | hrm, i'm not explaining this well. there's a few more points to mention | 18:35 |
| kanzure | 1) you can do this in continuous flow with an array of these reactions, but they have to all be growing the same sequence so that you don't contaminate different reactions | 18:35 |
| kanzure | 2) you theoretically want to do dna synthesis of different sequences in parallel; but you'd have to keep each reaction physically isolated | 18:36 |
| azonenberg | Hmm | 18:36 |
| kanzure | if you are doing this in continous flow with a shared reagent/wash bath for all beads (on which this reaction is occurring), | 18:36 |
| kanzure | hmm no. the point i am trying to make is that because of the error rate of this chemistry you want to monitor each bead and make sure the sequence is correct | 18:38 |
| azonenberg | Ooh, interesting | 18:38 |
| kanzure | if you are in a continous flow situation, how do you physically move an individual bead to lead it down the path to, say, sequencing | 18:38 |
| azonenberg | By raman spectroscopy? | 18:38 |
| azonenberg | (monitoring) | 18:38 |
| kanzure | well, that's one way, but i haven't seen a paper doing that yet | 18:38 |
| kanzure | the most basic version of all of this is don't use DLP/DMD, do only one oligonucleotide synthesis at a time on a single solid support bead, don't do anything parallel and hope for the best.. | 18:39 |
| kanzure | but this loses the advantages of microscopic features | 18:39 |
| kanzure | i mean why not just pick up a syringe and do it, at that point ;) | 18:39 |
| kanzure | syringe method: http://diyhpl.us/~bryan/papers2/DNA/Syringe%20method%20for%20stepwise%20chemical%20synthesis%20of%20oligonucleotides.pdf | 18:39 |
| azonenberg | Hmm, i'm not familiar enough with the techniques to suggest much at this point lol | 18:40 |
| kanzure | yeh i just need to rant to someone heh | 18:40 |
| kanzure | i mean, i guess it's not too disastrous if i have an unmonitored collection of a million beads simultaneously (but if it's unmonitored, and under continous flow, i don't need the photochemistry and all the sequences would (hopefully) be the same) | 18:41 |
| kanzure | and then hope that at least one of the beads has at least one strand of DNA that ends up being correct | 18:43 |
| azonenberg | Lol | 18:44 |
| azonenberg | How do you find it' | 18:44 |
| azonenberg | and separate it | 18:44 |
| kanzure | whether or not "finding it and separating it" is a problem depends on which method you're using | 18:46 |
| kanzure | for instance, if you are under continous flow where all beads get the same reagants/wash simultaneously, you hope for the best that all beads are synthesizing/growing the same sequence | 18:47 |
| kanzure | when you force the beads out of the system, you wouldn't care which one is which | 18:47 |
| kanzure | since they all theoretically are the same | 18:47 |
| azonenberg | But in practice? | 18:48 |
| kanzure | in practice- in that method- there are definitely going to be certain strands that are wrong | 18:48 |
| kanzure | oligonucleotide chemistry has error rates as bad as 1 in 250 bp wrong | 18:48 |
| kanzure | (enzymes like dna polymerase make 1 error in 10,000...) | 18:48 |
| kanzure | so what if you're synthesizing a 800 bp strand.. and the 750th bp is wrong. you're going to be pissed off. | 18:49 |
| azonenberg | Lol, yeah | 18:49 |
| azonenberg | Can you make say four 200bp strands separately | 18:49 |
| azonenberg | verify each | 18:49 |
| azonenberg | then concatenate? | 18:50 |
| azonenberg | kinda like Xilinx does with their virtex 7 chips | 18:50 |
| kanzure | yes you can concatenate | 18:50 |
| kanzure | this is called ligation | 18:50 |
| azonenberg | ok | 18:51 |
| kanzure | verification is its own can of worms | 18:51 |
| azonenberg | Is it worth doing to boost yieldss? | 18:51 |
| azonenberg | And the way i'd verify would be | 18:51 |
| kanzure | you can verify by dna sequencing on a microfluidic chip ;) | 18:51 |
| azonenberg | to PCR each of the test strands | 18:51 |
| azonenberg | then sequence some of it | 18:51 |
| kanzure | right | 18:52 |
| kanzure | pcr on a chip is really amazing.. my favorite method is LED-assisted heating of a droplet of water. it's like 20x faster than lab equipment since the volume is so small | 18:52 |
| azonenberg | very fast thermal cycling? | 18:52 |
| azonenberg | nice | 18:52 |
| kanzure | right.. | 18:53 |
| kanzure | http://diyhpl.us/~bryan/papers2/microfluidics/pcr/PCR%20-%20Nanodroplet%20real-time%20PCR%20system%20with%20laser%20assisted%20heating.pdf | 18:53 |
| kanzure | so, dna sequencing needs as much optimizing as dna synthesis, so it basically doubles the complexity of the project | 18:54 |
| kanzure | but dna sequencing is important to me so i'm willing to overlook that | 18:54 |
| azonenberg | i see | 18:55 |
| azonenberg | Yeah, i've never done full sequencing | 18:55 |
| azonenberg | only simple fingerprinting by gel electrophoresis | 18:55 |
| kanzure | you have to select what you are going to sequence. in a continous flow situation with DLP, how would you move beads out to individually test (or rather- how would you send the "detach from bead" chemistry to only that bead) | 18:56 |
| azonenberg | looking for presence or absence of one or two SNPs | 18:56 |
| kanzure | if i can't come up with a "access a specific bead under continuous flow" solution, then the DLP is more or less useless :P | 18:56 |
| azonenberg | Hmm, i see | 18:56 |
| kanzure | "just put 1000s of valves in your circuit and it'll be fine" | 18:57 |
| kanzure | one possibility is to use droplets instead (water-in-oil).. a set of beads per droplet. store each droplet at an xy location. the wash step would involve.. uh. | 18:58 |
| kanzure | i guess a droplet wash step would involve (1) a clean drop of water + (2) wash chemistry + (3) a way to dilute the bead/droplet's water enough times over? | 18:59 |
| azonenberg | Hmm | 19:18 |
| kanzure | my connection sucks today. | 19:28 |
| kanzure | i think i've found a good solution | 19:28 |
| kanzure | i'll store 100~ beads per droplet. water-in-oil microfluidics. | 19:29 |
| kanzure | store each droplet in a register specific to the next reaction that it needs to have. i.e., store all the "give this a G" together. | 19:29 |
| kanzure | move the droplets out of the register into a continuous flow channel. space them out by distance, so that 100 beads from droplet 1 are physically separated from 100 beads from droplet 2 | 19:30 |
| kanzure | magnetize the beads, do continuous flow microfluidics to do the synthesis cycle/steps, waste water output | 19:30 |
| kanzure | turn off waste; redirect flow to a feature that re-forms a droplet based on timing, so that the beads from the original droplets end up in separate droplets again | 19:31 |
| kanzure | depending on the length of the reaction channel maybe you'll react 100s of different sequences simultaneously | 19:31 |
| kanzure | then you cycle between doing A, C, T and G reactions in the continuous flow "reaction channel" to build up your final sequence, based on what you have loaded into the reaction channel at the moment | 19:32 |
| azonenberg | Hmm | 19:34 |
| azonenberg | So each of those 100 beads are considered equivalent? | 19:34 |
| kanzure | yeah; there can be any number (but hopefully at least one) growing oligo per bead | 19:35 |
| kanzure | maybe there's 10,000 strands of dna growing on a single bead | 19:36 |
| kanzure | but adding beads is nice for redundancy's sake | 19:36 |
| azonenberg | Ok | 19:37 |
| azonenberg | But you cant grab one bead separately from the others | 19:37 |
| azonenberg | only drop from drop | 19:37 |
| kanzure | yes, because of the spacing/timing when the other beads were released into the reaction channel | 19:39 |
| kanzure | i.e. release first droplet, let the droplet break up and go downstream for 10-20 ms (or whatever the bead formation time constant is...) | 19:39 |
| kanzure | *droplet formation | 19:39 |
| azonenberg | Yeah, makes sense | 19:40 |
| kanzure | in this scenario, you hope that each bead within the same droplet is growing the same oligo | 19:40 |
| kanzure | so if you want to sequence a droplet, you take just one bead out and go pcr it and do other things | 19:40 |
| azonenberg | And then keep the other beads going | 19:41 |
| azonenberg | interesting | 19:41 |
| azonenberg | how big are these beads? | 19:41 |
| azonenberg | tens of microns? | 19:41 |
| azonenberg | single microns? | 19:41 |
| kanzure | tons of companies sell tons of different beads | 19:42 |
| kanzure | look up polystyrene beads on google scholar | 19:42 |
| kanzure | but yes i've seen 10 micron beads, definitely | 19:42 |
| kanzure | interestingly enough there are microfluidic devices capable of manufacturing polystyrene beads as well... | 19:43 |
| azonenberg | what i meant was, more, what is a typical size | 19:44 |
| azonenberg | or size range | 19:44 |
| azonenberg | and are they porous or pretty solid | 19:44 |
| kanzure | dunno. 1 micron beads exist too. | 19:44 |
| kanzure | solid | 19:44 |
| azonenberg | And the DNA just sticks on the surface? | 19:44 |
| kanzure | there's some chemistry for that | 19:46 |
| kanzure | i'm really bad at magnetism, i don't know if a magnetic microbead could be kept in place with an electromagnet directly under it, or if the flow forces would sweep it away | 19:48 |
| azonenberg | Depends on the conditions, there's no definite yes or no | 19:49 |
| azonenberg | depends on cross section, flow velocity, amoutn of magnetic mateiral in the bead, strength of the electromagnet's field | 19:49 |
| kanzure | flow velocity can be varied by me, so that's not a relevant variable | 19:50 |
| kanzure | actually, if the overall flow velocity is low enough, i bet a bead couldn't be moved | 19:50 |
| azonenberg | Well, in that case i'd say it can definitely be done | 19:50 |
| azonenberg | the question is how strong a magnet will you need | 19:50 |
| azonenberg | and how magnetic you can make the bead | 19:50 |
| kanzure | they wouldn't have to be magnetic if i'm ok with very low velocities | 19:51 |
| kanzure | and then increase the flow rate when i want to move them | 19:51 |
| kanzure | ah but that would make the beads closer to the "top" move first, sadly | 19:51 |
| kanzure | hi gene_hacker | 19:54 |
| kanzure | gene_hacker: azonenberg is around now, you had some questions.. | 19:54 |
| gene_hacker | is azonenberg here right now? | 20:03 |
| azonenberg | lol, missed him again | 21:42 |
| --- Wed Feb 1 2012 | 00:00 | |
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