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A Visual Guide to DNA Sequencing

58 points by surprisetalk 8mo ago

jonn_smith 8mo ago
Nice writeup, but missing 2 newer sequencers: Roche's Sequencing By Expansion platform and UltimaGenomics' platform (edited for clarity)
- mfld 8mo ago
  Yes, they are only mentioned in the last section. Other notable misses would be Ion Torrent and MGI. Still a nice article with a focus on the key technologies.
voidUpdate 8mo ago
So it's not just a strong magnifying glass and tiny tweezers?
- adrian_b 8mo ago
  The nanopore technique described towards the end is not that far from your description.
- dnautics 8mo ago
  they kinda tried that and it is my understanding that it sucks. One example of "direct linear mapping": https://pdfs.semanticscholar.org/5080/78a97f4778eee488e11032039fedb985a36e.pdf https://pdfs.semanticscholar.org/5080/78a97f4778eee488e11032...
ben1040 8mo ago
I worked in this space for over a decade. It's so weird to me to see the workhorse sequencing hardware like the ABI 37x/37xx series, 454, and Illumina machines now as literal museum pieces.
- searine 8mo ago
  Agreed. Seeing million dollar sequencers on ebay for a few thousand bucks makes me want to reach for my wallet, but the I realize there are no reagents for it anyway.
  - ben1040 8mo ago
    Back in 1999 I joked that the ABI 3700 looked like a $300K mini fridge. Now you could actually get one on eBay and convert one for cheap.
  - rootsudo 8mo ago
    Can you expand more on this? I’ve seen said machines too but I assumed you could use different solutions and just have a contract with sigma Aldrich to get what’s needed - what else would someone have to do to put a used sequencer to good use?
    - ben1040 8mo ago
      There are more consumables involved there beyond just reagents and a lot of it was/is proprietary. The machines all had their own consumable physical bits for sequencing as well and getting your hands on those would also be a blocker. Applied Biosystems Sanger capillary machines (the 37XX series) had polymers that were injected into the capillaries to reflow between runs (the polymer took the place of the physical slab of gel). The capillaries themselves were also consumable and would wear out after so many runs. They were extremely expensive even 20-30 years ago when these were state of the art. You also need the dideoxy dye terminators you read about in the article. The patents on BigDye are expired now (the patents were ABI's principal moat on the Sanger tech and they aggressively used it against competitors like MegaBACE) but it's not like that's a growth market for a vendor trying to make a generic replacement. Similar with the 454 -- the process would fragment up the DNA into lots of tiny shards, and bind the fragments to microscopic beads. There was a physical flow cell chip that you would flow the beads onto. The chip had microscopic "beehive" cells to hold the beads such that you had a vast array of miniature test tubes running reactions in parallel while the machine imaged the result. These chips were one and done after every sequencer run, and were precision manufactured. They were manufactured by fusing together a bundle of fiber optics and then slicing it like bread, such that you had a glass chip and the space between the fibers became the cells. You'd need the beads, the chips, and the reagents to produce the reactions the machine is looking for. Without all the reagents and physical necessities these machines are just some really fancy cameras with onboard lasers. Long story short it's fun to think about getting one of these machines, but unless you use it as a cool looking coffee table that appeals to graying molecular biologists, you're going to be disappointed.
urza 8mo ago
We need PC revolution for biology. Put a box on every table we can tinker with.
- conception 8mo ago
  You can get a USB sequencer for about 150 bucks
- OkayPhysicist 8mo ago
  The equipment necessary is broadly within the range that early PCs were. There's just a lot fewer biohackers than there were computer hackers.
joshuamcginnis 8mo ago
I culture fungus, perform DNA extraction, run PCR (polymerase chain reaction - the process used to clone the gene(s) I want to sequence), and gel electrophoresis in my home lab. I then ship tubes of the DNA to mclab.com where for about $3 per sample, they'll run sanger sequencing and email me the data the next day. Works great for DNA barcoding and plasmid sequencing.

58 points by surprisetalk 8mo ago

Nice writeup, but missing 2 newer sequencers: Roche's Sequencing By Expansion platform and UltimaGenomics' platform (edited for clarity)

mfld 8mo ago
Yes, they are only mentioned in the last section. Other notable misses would be Ion Torrent and MGI. Still a nice article with a focus on the key technologies.

voidUpdate 8mo ago

So it's not just a strong magnifying glass and tiny tweezers?

adrian_b 8mo ago
The nanopore technique described towards the end is not that far from your description.
dnautics 8mo ago
they kinda tried that and it is my understanding that it sucks. One example of "direct linear mapping": https://pdfs.semanticscholar.org/5080/78a97f4778eee488e11032039fedb985a36e.pdf https://pdfs.semanticscholar.org/5080/78a97f4778eee488e11032...

ben1040 8mo ago

I worked in this space for over a decade. It's so weird to me to see the workhorse sequencing hardware like the ABI 37x/37xx series, 454, and Illumina machines now as literal museum pieces.

searine 8mo ago
Agreed. Seeing million dollar sequencers on ebay for a few thousand bucks makes me want to reach for my wallet, but the I realize there are no reagents for it anyway.
- ben1040 8mo ago
  Back in 1999 I joked that the ABI 3700 looked like a $300K mini fridge. Now you could actually get one on eBay and convert one for cheap.
- rootsudo 8mo ago
  Can you expand more on this? I’ve seen said machines too but I assumed you could use different solutions and just have a contract with sigma Aldrich to get what’s needed - what else would someone have to do to put a used sequencer to good use?
  - ben1040 8mo ago
    There are more consumables involved there beyond just reagents and a lot of it was/is proprietary. The machines all had their own consumable physical bits for sequencing as well and getting your hands on those would also be a blocker. Applied Biosystems Sanger capillary machines (the 37XX series) had polymers that were injected into the capillaries to reflow between runs (the polymer took the place of the physical slab of gel). The capillaries themselves were also consumable and would wear out after so many runs. They were extremely expensive even 20-30 years ago when these were state of the art. You also need the dideoxy dye terminators you read about in the article. The patents on BigDye are expired now (the patents were ABI's principal moat on the Sanger tech and they aggressively used it against competitors like MegaBACE) but it's not like that's a growth market for a vendor trying to make a generic replacement. Similar with the 454 -- the process would fragment up the DNA into lots of tiny shards, and bind the fragments to microscopic beads. There was a physical flow cell chip that you would flow the beads onto. The chip had microscopic "beehive" cells to hold the beads such that you had a vast array of miniature test tubes running reactions in parallel while the machine imaged the result. These chips were one and done after every sequencer run, and were precision manufactured. They were manufactured by fusing together a bundle of fiber optics and then slicing it like bread, such that you had a glass chip and the space between the fibers became the cells. You'd need the beads, the chips, and the reagents to produce the reactions the machine is looking for. Without all the reagents and physical necessities these machines are just some really fancy cameras with onboard lasers. Long story short it's fun to think about getting one of these machines, but unless you use it as a cool looking coffee table that appeals to graying molecular biologists, you're going to be disappointed.

urza 8mo ago

We need PC revolution for biology. Put a box on every table we can tinker with.

conception 8mo ago
You can get a USB sequencer for about 150 bucks
OkayPhysicist 8mo ago
The equipment necessary is broadly within the range that early PCs were. There's just a lot fewer biohackers than there were computer hackers.

joshuamcginnis 8mo ago

I culture fungus, perform DNA extraction, run PCR (polymerase chain reaction - the process used to clone the gene(s) I want to sequence), and gel electrophoresis in my home lab. I then ship tubes of the DNA to mclab.com where for about $3 per sample, they'll run sanger sequencing and email me the data the next day. Works great for DNA barcoding and plasmid sequencing.