--- Log opened Fri Feb 12 00:00:27 2021 00:03 -!- thahxa_ [~thahxa@vlnsm4-toronto63-142-122-137-196.internet.virginmobile.ca] has joined ##hplusroadmap 00:04 -!- thahxa [~thahxa@vlnsm4-toronto63-142-122-137-94.internet.virginmobile.ca] has quit [Ping timeout: 240 seconds] 00:16 -!- join_cordblood [~join_cord@135-23-248-163.cpe.pppoe.ca] has joined ##hplusroadmap 00:26 -!- Llamamoe [~Llamagedd@088156213212.radom.vectranet.pl] has joined ##hplusroadmap 00:33 < thahxa_> does anyone know the specifics of how liquids are transported into the columns for oligo synthesis? 00:33 -!- thahxa_ is now known as thahxa 00:34 < thahxa> like is it injected from the top? bottom? how does it flow out? 00:34 < thahxa> ive been searching for a while (but maybe inthe wrong places?) and i cant seem to get an answer 00:37 < L29Ah> the synthesizer i've seen injects stuff with a few needles at the top and has a tube at the bottom of each reactor 00:38 < thahxa> few needles per column? 00:38 < thahxa> or is it just one needle that gets flushed with mecn between runs 00:40 < thahxa> but yea that was what i was semi-suspecting 00:40 < L29Ah> thahxa: the needles are located on a 3d robot 00:41 < thahxa> so they inject /into/ the column? 00:41 < L29Ah> so the device can do a shitload of tiny syntheses simultaneously 00:41 < L29Ah> kinda 00:41 < thahxa> wait what model is this? 00:41 < fenn> the trolltron 9000 00:42 < L29Ah> idk some half-assed old Gilson stuff from ebay 00:42 < thahxa> or more rather it does column synthesis right? 00:42 < thahxa> ah ok 00:42 < L29Ah> the synthesis occurs on some fritty shit on a resin afaiu 00:43 < L29Ah> er, the other way around 00:43 < thahxa> yeah that part im familiar with 00:43 < thahxa> the question was really how the columns work with insertion/drainage 00:43 < L29Ah> imagine the middle piece of a three-piece single-use syringe 00:44 < L29Ah> only with a frit 00:44 < thahxa> middle piece as in the barrel? 00:44 < L29Ah> yes 00:44 < thahxa> alright thx 00:44 < fenn> so it's luer lock? 00:45 < fenn> luer taper 00:45 < L29Ah> i think there weren't any fancy joints 00:45 < thahxa> hm alright then 00:45 < fenn> see pg 30 http://tools.thermofisher.com/content/sfs/manuals/cms_095580.pdf 00:46 < thahxa> time to stuff this information in my head until i can do somethin wtih it lmao 00:47 < fenn> also pg 68 00:47 < thahxa> ah alright i see now 00:48 < fenn> i don't remember if it was you asking about reagent costs, but you can buy a 500ml bottle of each base for less than $100 00:48 < thahxa> i was 00:48 < thahxa> my idea is that its probably cheaper to buy straight from alibaba 00:48 < thahxa> and mix it yourself 00:48 < fenn> well you don't need a kilogram of each base 00:48 < thahxa> nah i wasnt asking for a kilo 00:48 < thahxa> only 25g of each 00:48 < L29Ah> the solvents are a PITA, they dissolve every fucking thing 00:49 < thahxa> its just mecn right? 00:49 < fenn> acetonitrile and THF 00:49 < thahxa> i have it on good authority (i googled it) that pp is mecn resistant 00:49 < thahxa> oh yea thf is probably gonna b an issue lmao 00:49 < thahxa> (not to mention the uh peroxide problem) 00:51 < thahxa> fenn: wait where can you buy the bases for that cheap? 00:51 < thahxa> and at what concentration? 00:51 < fenn> an online store that sold used synthesizers which i can't remember the name of 00:52 < thahxa> ah sad 00:52 < thahxa> cuz thats a pretty good price 00:52 < thahxa> wait 00:52 < thahxa> is it for /all/ the bases? 00:52 < fenn> i don't remember 00:52 < thahxa> mk ill look into it then 01:02 < fenn> https://azcobiotech.com/reagents-for-oligonucleotide-synthesis/phosphoramidites/ might be in the wayback machine 01:08 < fenn> aww javascript strikes again 01:10 < fenn> is "5' chemical phosphorylation agent" what you want? it was $150/g 01:10 < thahxa> uh no 01:11 < thahxa> the price was for something like DMT-dA(bz) 01:11 < thahxa> or similar 01:12 < fenn> "5' dimethoxytrityl N-acetyl 2'deoxyCytidine3'[(2-cyanoethyl)-(N-N-diisopropyl)]-phosphoramidite"? $6/g 01:13 < thahxa> ah thx 01:13 < thahxa> thats slightly more expensive than the china price i got quoted 01:13 < fenn> http://web.archive.org/web/2011*/azcobiotech.com:80/reagents/7.htm 01:13 < fenn> that's the 1 gram price 01:14 < fenn> you might have better luck poking around here than me http://web.archive.org/web/2011*/azcobiotech.com:80/reagents/* 01:14 < thahxa> ah thanks 01:14 < thahxa> i was really just asking to probe 01:15 < thahxa> as an introdcutory "how much /should/ this cost" sorta thing 01:15 < thahxa> well the chinese sellers anyways 01:15 < fenn> right 01:15 < thahxa> just to get a grasp of how expensive all this stuff is (answer: pretty damn expensive) 01:15 < fenn> i don't know what happened to azcobiotech but their website disappeared around 2016 01:15 < thahxa> yea 01:15 < thahxa> probably went bankrupt or somethin 01:15 < thahxa> i think i got highballed on their dci offers tho 01:15 < fenn> well for DNA you don't need more than a few pmol because you can amplify it with polymerase 01:16 < thahxa> in a sense yea but i cant find any columns smaller than 40nmol 01:16 < thahxa> and even then you need a 20-fold excess every step iirc 01:16 < thahxa> and im a bit iffy about the printer route 01:16 < thahxa> imo 01:17 < fenn> these big clunky column synthesizers are for making PCR primers 01:17 < thahxa> really? 01:17 < fenn> for that you actually do need a relatively large quantity of DNA 01:17 < thahxa> ok so some background is that im more used to chemistry scales 01:17 < thahxa> so EVERYTHING in bio looks small scale to me 01:18 < thahxa> which is probably why im running around like an idiot asking for tons of reagents 01:18 < fenn> supposedly they have less error but it works out to like 200 bp oligos instead of 100 bp oligos 01:18 < fenn> so you might as well just make 1000x more in parallel with an optical or inkjet system 01:18 < thahxa> hm true 01:18 < fenn> then deal with the error later when combining the shorter oligos 01:18 < thahxa> wait how are you supposed to detach from column in an inkjet system anyways? 01:19 < thahxa> oh and the second reason why i was leaning towards column was simplicity 01:19 < fenn> lol simplicity is not having a million tubes and valves 01:19 < thahxa> my idea was just to go full on with syringes 01:19 < fenn> there's some enzyme that cuts the first few bases which are special, i don't remember exactly how it works 01:19 < thahxa> like schlenk line chem 01:20 < thahxa> ah 01:20 < fenn> or you can encode "cut here" in the first few bases you write 01:20 < thahxa> restriction enzymes are the name btw 01:20 < fenn> right 01:20 < thahxa> but yea my idea was that the machine cost was way higher than any of the chemical costs 01:21 < fenn> i thought there was a scheme where the linker is chemically different and comes from the factory pre-seeded with reaction sites 01:21 < thahxa> yes 01:21 < thahxa> they use a succinate bond (butanedioic acid) 01:21 < thahxa> wait actually 01:21 < thahxa> somethign i never asked 01:21 < thahxa> but whats the typical level of organic chem experience around here? 01:22 < fenn> uh.. none is typical 01:22 < thahxa> that explains a few things here 01:22 < thahxa> well it is a bio community after all 01:22 < fenn> i took freshman organic lab in college which was punishment more than anything 01:23 < thahxa> lol 01:23 < fenn> it's just taught so badly.. all memorization and no understanding 01:23 < thahxa> tbf my ochem knowledge isnt much better 01:24 < thahxa> either way i was wonderin about a scheme to "compound" long oligos together chemically 01:24 < thahxa> has anyone tried that yet? 01:24 < fenn> you need to read about biotech 01:24 < fenn> also there's a huge paradigm shift between ochem and bio 01:24 < fenn> despite being made out of the same stuff 01:25 < thahxa> ? in what sense 01:25 < fenn> bio is so much more precise and produces no side products 01:25 < thahxa> ah 01:26 < thahxa> in a sense maybe 01:26 < thahxa> actually probably yea 01:27 < thahxa> either way i was just wonderin if anyone has actually tried sticking two long oligos together with chem 01:27 < fenn> anyway yes there is an enzyme called ligase that automatically makes the covalent bonds when two pieces of DNA are stuck together with hydrogen bonds. there's also another type of ligase that randomly combines blunt ended cuts (no overlapping single stranded DNA) 01:27 < thahxa> yea ik about ligase 01:27 < fenn> don't understand the question then 01:27 < thahxa> issue is that with blunt ends its not very selective right? 01:27 < fenn> right 01:27 < thahxa> so my idea was just to grab the oligo 01:28 < thahxa> stick a dmt group on the top and a phosphocyanowhatever group on the bottom 01:28 < thahxa> and stick them together like in the traditional dna synthesis 01:28 < fenn> but why 01:28 < thahxa> i was just wondering if anyones tried it, and why it (probably) doesnt work 01:29 < thahxa> the "but why" is basically just probably better selectivity compared to ligaseing everythign together and hopefully more flexibility? 01:29 < fenn> the idea is you only get one possible combination of oligos? 01:29 < thahxa> yes 01:31 < fenn> well i don't know what's involved in "just stick a DMT group on it" 01:31 < thahxa> its reaction with dimethoxytrityl chloride 01:31 < thahxa> actually 01:31 < thahxa> the ideal was to make one oligo dmt-on 01:31 < thahxa> so you wouldn't have to remove it at all 01:31 < fenn> presumably there are side reactions where you get DMT groups in the wrong place, and those could cause mutations later on when replicating the DNA 01:32 < thahxa> yea the issue is that i cant find much research on it 01:32 < thahxa> part of the idea is based on an oldish book 01:32 < fenn> so you add a special base at the end? 01:32 < fenn> during oligo synthesis 01:33 < thahxa> you dont need to because you can make oligos dmt-on so you dont need anything special at the end 01:33 < fenn> no uncapping step at the end then 01:33 < thahxa> and the idea would be to stick a thing at the (uh 3'-end i think it is?) and to couple chemically from there 01:33 < thahxa> yes 01:34 < fenn> then the problem is putting the phosphocyanowhatever group on 01:34 < thahxa> (the stick onto 3'end part can be done in about 70-80% yield according to that book apparently, and i think you can use pcr amplificatino for part of it maybe? but that seems a bit more complicated) 01:34 < thahxa> from what i read its not too difficult, but ive never heard of it being done ona whole oligo 01:34 < thahxa> (although im sure brute force probably works here) 01:49 * fenn points any time travelers at http://gnusha.org/logs/2015-07-13.log 01:50 < thahxa> huh nice 01:50 < thahxa> oh and the source of the "idea" is "Protocols for Oligonucleotides and Analogs Synthesis and Properties" 01:51 < thahxa> on page 44 (section 3.5) 01:51 < thahxa> ill note that you can probably sub out tetrazole for something nicer 01:52 < fenn> if your goal is to just jam two oligos together and get a known product out, you can make them different lengths and run it through a gel or column to select for length 01:52 < fenn> er, with ligase 01:52 < thahxa> true that actually 01:52 < thahxa> didnt think of that 01:52 < fenn> the multiple additions or self-additions will get filtered out 01:52 < fenn> you can actually do several ligase reactions at once that way 01:53 < thahxa> although there would be an issue of joining AB or BA 01:53 < fenn> it gets exponentially more difficult with more serial additions in the same pot 01:53 < thahxa> (i guess you'd have to do some capping steps or something?) 01:54 < fenn> the single strand overhang with restriction sites at the joints is how it's usually done now 01:54 < fenn> so you end up with a final product without the restriction sites in it 01:54 < thahxa> yeah i read about that 01:55 < thahxa> seems a bit limited though 01:55 < thahxa> although im sure its manageable 01:55 < fenn> there are a few different schemes so you can pick the appropriate scheme if you just have to have that restriction sequence in there 01:55 < fenn> but usually you can swap a codon or something 01:55 < thahxa> yeah 01:56 < thahxa> ig the ideal would be to have a restriciton enzyme that cuts like GGGGGG or GCGGCG or somethin 01:56 < thahxa> so you can just put gly or ala on those sites 02:15 < thahxa> actually truly idealy would be some magic that can automatically select which oligos to make and which restriction enzymes to put where when you give it the amino acid sequence 02:22 -!- thahxa [~thahxa@vlnsm4-toronto63-142-122-137-196.internet.virginmobile.ca] has quit [Ping timeout: 264 seconds] 02:39 -!- darsie [~kvirc@84-113-55-200.cable.dynamic.surfer.at] has joined ##hplusroadmap 03:30 -!- midnight [~midnight@unaffiliated/midnightmagic] has quit [Ping timeout: 240 seconds] 03:33 -!- midnight [~midnight@unaffiliated/midnightmagic] has joined ##hplusroadmap 04:29 -!- sivoais_ [~zaki@199.19.225.239] has joined ##hplusroadmap 04:31 -!- sivoais [~zaki@unaffiliated/sivoais] has quit [Ping timeout: 240 seconds] 05:01 -!- sanehatter_ [sanehatter@gateway/vpn/mullvad/sanehatter] has joined ##hplusroadmap 05:03 -!- sanehatter [sanehatter@gateway/vpn/mullvad/sanehatter] has quit [Ping timeout: 240 seconds] 06:08 -!- balrog [~balrog@unaffiliated/balrog] has quit [Ping timeout: 240 seconds] 06:09 -!- balrog [~balrog@unaffiliated/balrog] has joined ##hplusroadmap 06:42 -!- spaceangel [~spaceange@ip-94-112-205-34.net.upcbroadband.cz] has joined ##hplusroadmap 06:53 -!- sanehatter_ [sanehatter@gateway/vpn/mullvad/sanehatter] has quit [Quit: -] 06:58 -!- juri_ [~juri@178.63.35.222] has quit [Ping timeout: 256 seconds] 06:59 < gnusha> https://secure.diyhpl.us/cgit/diyhpluswiki/commit/?id=a0d813e6 Michael Folkson: Add Matt on TFTC >> http://diyhpl.us/diyhpluswiki/transcripts/tftc-podcast/2021-02-11-matt-corallo-taproot-activation/ 06:59 < gnusha> https://secure.diyhpl.us/cgit/diyhpluswiki/commit/?id=0ac4dace Michael Folkson: Merge pull request #206 from michaelfolkson/matt-activation-tftc >> http://diyhpl.us/diyhpluswiki/ 07:05 < L29Ah> huh, oligonucleotide synthesis as a service is pretty cheap 07:05 < L29Ah> even in my shithole 07:20 -!- juri_ [~juri@178.63.35.222] has joined ##hplusroadmap 07:36 < L29Ah> i wonder though how hard would it be to put it into an actual virus 08:59 -!- sanehatter [~sanehatte@141.98.255.147] has joined ##hplusroadmap 09:35 < superkuh> In the first improvement to google search in years it now shows DOI for some search results. 09:38 -!- thahxa [~thahxa@vlnsm4-toronto63-142-122-137-196.internet.virginmobile.ca] has joined ##hplusroadmap 10:38 -!- helleshin [~talinck@cpe-174-103-159-2.cinci.res.rr.com] has quit [Read error: Connection reset by peer] 11:07 -!- yashgaroth [~ffffffff@2601:5c4:c780:6aa0:4fc:c150:17e9:4254] has joined ##hplusroadmap 11:19 -!- filipepe_ [uid362247@gateway/web/irccloud.com/x-tkzwqwjyjuwfamzk] has joined ##hplusroadmap 12:34 -!- CryptoDavid [uid14990@gateway/web/irccloud.com/x-cqspboylyjvcheaz] has joined ##hplusroadmap 13:16 -!- Llamamoe [~Llamagedd@088156213212.radom.vectranet.pl] has quit [Quit: Leaving.] 15:48 -!- filipepe_ [uid362247@gateway/web/irccloud.com/x-tkzwqwjyjuwfamzk] has quit [Quit: Connection closed for inactivity] 15:51 -!- Cory [~Cory@unaffiliated/cory] has quit [Ping timeout: 264 seconds] 15:58 -!- join_cordblood [~join_cord@135-23-248-163.cpe.pppoe.ca] has quit [Ping timeout: 240 seconds] 15:59 -!- Cory [~Cory@unaffiliated/cory] has joined ##hplusroadmap 16:10 -!- join_cordblood [~join_cord@135-23-248-163.cpe.pppoe.ca] has joined ##hplusroadmap 17:05 -!- CryptoDavid [uid14990@gateway/web/irccloud.com/x-cqspboylyjvcheaz] has quit [Quit: Connection closed for inactivity] 17:12 -!- yashgaroth [~ffffffff@2601:5c4:c780:6aa0:4fc:c150:17e9:4254] has quit [Quit: Leaving] 17:22 -!- filipepe_ [uid362247@gateway/web/irccloud.com/x-ykigcmfkucmkjqwt] has joined ##hplusroadmap 17:23 -!- Codaraxis [Codaraxis@gateway/vpn/mullvad/codaraxis] has joined ##hplusroadmap 17:26 -!- Codaraxis_ [Codaraxis@gateway/vpn/mullvad/codaraxis] has quit [Ping timeout: 264 seconds] 17:41 -!- preview [~quassel@2407:7000:8423:b67:10cc:dd04:c0bf:daaf] has quit [Ping timeout: 240 seconds] 17:47 -!- preview [~quassel@210.246.47.5] has joined ##hplusroadmap 17:54 -!- justanotheruser [~justanoth@unaffiliated/justanotheruser] has quit [Ping timeout: 272 seconds] 18:08 -!- spaceangel [~spaceange@ip-94-112-205-34.net.upcbroadband.cz] has quit [Remote host closed the connection] 18:52 -!- Sir_Alexei [uid348072@gateway/web/irccloud.com/x-xzaozcoezcvvlcey] has joined ##hplusroadmap 19:21 < kanzure> .title https://news.ycombinator.com/item?id=26118571 19:21 < saxo> FIDO2 security key company releases hardware that's open source and uses Rust | Hacker News 19:21 < kanzure> https://solokeys.com/blogs/news/our-solo-v2-campaign-launches-on-january-26th 19:22 < kanzure> https://littlemaninmyhead.wordpress.com/2015/09/28/so-you-want-to-learn-to-break-ciphers/ 19:41 -!- filipepe_ [uid362247@gateway/web/irccloud.com/x-ykigcmfkucmkjqwt] has quit [Quit: Connection closed for inactivity] 19:51 -!- darsie [~kvirc@84-113-55-200.cable.dynamic.surfer.at] has quit [Quit: bye bye] 19:58 -!- Sir_Alexei [uid348072@gateway/web/irccloud.com/x-xzaozcoezcvvlcey] has quit [] 20:23 -!- preview [~quassel@210.246.47.5] has quit [Ping timeout: 240 seconds] 20:30 -!- filipepe_ [uid362247@gateway/web/irccloud.com/x-nqfxqrqgrlzohczu] has joined ##hplusroadmap 21:32 -!- thahxa [~thahxa@vlnsm4-toronto63-142-122-137-196.internet.virginmobile.ca] has quit [Remote host closed the connection] 23:09 -!- filipepe_ [uid362247@gateway/web/irccloud.com/x-nqfxqrqgrlzohczu] has quit [Quit: Connection closed for inactivity] 23:11 < L29Ah> .t https://www.nature.com/articles/s42005-021-00532-4 23:11 < saxo> Micro and nanoscale 3D printing using optical pickup unit from a gaming console | Communications Physics 23:35 -!- preview [~quassel@2407:7000:8423:b00::2] has joined ##hplusroadmap --- Log closed Sat Feb 13 00:00:27 2021