How We Got The DNA From This Extremely Rare Ciliate

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To study organisms at the genetic level, we need their DNA. Which means that we need to be able to wade through all the bits and pieces lying within their tiny bodies to pick out something even tinier-something we can’t just dig out with a shovel. So how does James manage to get the precious DNA from Legendrea loyezae and the other ciliates he’s interested in studying?
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4 дек 2022

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Комментарии : 184

@ke9tv Год назад

When I saw the previous episode on Legentrea, I thought, "I wonder if James has managed to isolate enough DNA from these to run a PCR and sequence." When I saw the title of this one, I cheered! PCR stands for "pipette, cry and repeat!"

@keku52 Год назад

lmfao, accurate acronym. My classmates (bachelors) trying to isolate STEC (Shiga toxin producing E. coli) and MRSA (Methicillin-Resistant S. aureus) have been crying how many times they have to repeat many times to extract enough genetic material for amplification and confirmatiin

@StopItGarrison 10 месяцев назад

Me too on being a gay lil dandy. I am very gay and will always be that way 😅

@Srcsqwrn Год назад

I'm only a little sad the little cilliate didn't gete to live out it's life. But the discovery was amazing, and being able to find it's siblings in genes is amazing!

@gtbkts Год назад

Same

@janetf23 Год назад

Same here too

@eg-fi3ju Год назад

you sound like a vegan it dosent have a brain its literally a single cell thats like being sad that your skin dies

@Srcsqwrn Год назад

@@eg-fi3ju ok boomer

@ivy_47 Год назад

@@eg-fi3ju what are your thoughts on pro-life philosophy?

@ColinTimmins Год назад

That "little guy" really stands out if you know how to look. Thank you for giving us all a look at the things we would have surely missed, every day, right under our nose.

@dianagibbs3550 Год назад

I remember being in a microbiology class in 1996ish and hearing about this new technology PCR, and how it was going to change the world. It has, and continues to do so. So cool.

@Bryophytan Год назад

These days it seems like CRISPR/cas9 is going to be an equivalent, if in a field skewed in a slightly different direction

@dianagibbs3550 Год назад

@@Bryophytan I was thinking that too!

@jimmyjames2022 Год назад

Congratulations to James and the research team for their work on this rare Legendrea ciliate, one of the coolest ciliates ever!

@osmia Год назад

James, you have such a fascinating job!

@TheTm9090 Год назад

Such humble beginnings and now we are making videos covering them. Absolutely mind blowing

@1234j Год назад

Well DONE, James! And your colleagues. Thank you for tracking the huge yet tiny stars and filming them for us, and thank you, all those on this channel who serve us up fresh knowledge and insights, one drop at a time - sometimes via the freezer. Cheers from England.

@obieobrien5883 Год назад

James must’ve been super excited to find this one!

@glennk.7348 Год назад

Can you also add the speed of filing to the data on screen? I was amazed to watch your 800x slower video and wondered how you even track some of these animals!

@LunarForte Год назад

They're all in real-time unless otherwise stated

@alfredwaldo6079 Год назад

This is truly a journey to the microscpe moment

@Syntaxxed Год назад

Thank you all so much. I will be doing a bachelor of physics or math. I really like that I can still be updated on biology and chemistry through channels like these.

@ppartsx Год назад

James explained to me months ago on r/microscopy how he gets the dna from singular microbes so i had an idea already how it was done before watching :]

@speckledjim_ Год назад

Absolutely stunning footage, so crisp and clear. i cant help but think that this little guy is similar in appearance to a cuttlefish or squid

@Bryophytan Год назад

It's the quality you get with ridiculous expensive microscopes.

@MeetAnEcoregion Год назад

7:58 Looking at old research with new eyes. I like that notion very much.

@WAMTAT Год назад

Amazing work James.

@THELITTLERIVERNERD Год назад

Love this video! Would love more videos that dig into the biochemistry & genetics of the microcosmos!

@rot_studios Год назад

Wondered about this lately, thank you so much for making a video about it

@cantsay Год назад

Thank you for sharing James amazing journey with us. What an amazing discovery, I'm sure his mind it blown, and will keep exoding in his head for months.

@FBIandre123 Год назад

As a portuguese watcher and lover of the calm and charismatic voice of hank green, is a little bit annoying having to change the voice in begining of every video.

@whizthesugoi Год назад

It's an awesome feature though

@KitsuNoir Год назад

RU-vid loves telling you what you prefer. I love when they choose a random language when English is unavailable for captions. Like, no, just because there's no English subs doesn't mean i can read Spanish or German suddenly, just turn it off till i see another video. Developers, don't assume stupid things, like that everyone speaks the same language or can only speak one. Or assume we can only speak one instead of simultaneously assuming contradictory ideas.

@1234j Год назад

Just hop forward a few seconds when you start the video, so you get all the soothing and none of the normal - just don't tell Hank.

@whizthesugoi Год назад

@@1234j no no, they mean the auto translate audio feature that is auto selected if you're from a portuguese or spanish speaking country

@belstar1128 Год назад

They just use text to speech i tried it in Portuguese because i am learning Portuguese. but i can't stand the text to speech voice i thought they would have a system where they can have multiple soundtracks for different languages. but with an actual human voice over but i will sick to the original for now.

@Streetsy Год назад

This channel is really interesting. Love your work.

@basedlordprime Год назад

Very cool to see it swimming in the water drop.

@mineovernutter Год назад

Can you add James' paper to the list of sources in the description? I think it is important to put it in there.

@nefeli2737 Год назад

Good episode. I'm in search of this microbe now.

@you2angel1 Год назад

WOW °~.🌟.~° You guys should be quite proud!

@trevinbeattie4888 Год назад

The thought of a tiny microbe taking a bath amuses me. :D

@WAMTAT Год назад

Not often you watch a RU-vid video on the cutting edge of science.

@vernonbrechin4207 Год назад

Thank you for answering some of the pre-PCR preparation questions that I had. I admire the patience it takes to engage in all those repetitive processes, especially that of cleaning the minute organism. Beyond that step I wonder why the DNA code is not damaged, or scrambled, by the physically disruptive steps that follow the 'washing.' Is the organism washed in distilled water, or water containing some nutrient minerals? It seems to me that even this washing step would not rid the subject organism of parasitic organisms that might be in it, such as viruses.

@TheRedKnight101 Год назад

James looked specifically at the 18S ribosomal RNA sequence which is only in eukaryotes. Only that sequence was amplified so any DNA from bacteria, archaea, and viruses would not be amplified. That is why it was important for the cell to degrade any DNA from other ciliates it may have eaten because that DNA could be amplified if they weren't careful.

@vernonbrechin4207 Год назад

@@TheRedKnight101 Thank you for your clear and thorough explanation.

@HesderOleh Год назад

@@TheRedKnight101 I figured that it was 18S barcoding based on the way it was described, which made me wonder why not get the whole genome?

@Raydensheraj Год назад

@@TheRedKnight101 The Carl Woese method....what a freaking genius the crazy old man was....

@dixietenbroeck8717 Год назад

*Poor little critter!* At least we can say "it devoted it's life to science" - _SOLELY_ for the benefit of a *COMPARATIVELY **_ENORMOUS,_** (and UNKNOWN) **_ORGANISM!_*

@ronaldhowell2005 Год назад

Thankyou for sharing such important information and video, awesomeness !!!

@Taomantom Год назад

always learn something new here thanks!

@DracarmenWinterspring Год назад

I wonder, for such a rare find, is it worth trying to give it the time and environment to replicate, if only to have a "backup" in case the DNA extraction fails? Or is there too much of a risk of it dying or losing track of it before it has a chance to reproduce?

@michable100 Год назад

Well it will not have been alone in the environment it was found in. So if you mess up you can always go back and try to catch more.

@Mei-oe6sg Год назад

I’m so appreciate these videos, thanks a lot 🙏 ❤❤

@SeahamV2 Год назад

I have to come back to these videos, as its information overload for me. Always amazing though!!!

@shgstewart4674 Год назад

It's a rather pretty little organism. Nicely done!

@bugpal Год назад

Do you amazing people have any plans or ideas for a documentary length feature that can be shown at IMAX Theatres, for example? That would truly be a sight to see.

@carsonclayson7063 Год назад

It just hit me that vibe-wise Journey to the microcosmos is Hank's Anthropocene reviewed

@ptarmigan1356 Год назад

Can I ask how James decided to use freeze thaw to lyse the cells? How did he know how many cycles would be enough? For comparison I have heard of using lithium acetate and SDS to lyse yeasts, followed by precipitation of DNA by ethanol. Also what is the structure of these organisms like, eg do you know if they have a cell wall? How different are different ciliates, what sort of things could James assume about these organisms before knowing from sequence data which family they belong to?

@HesderOleh Год назад

One reason to use freeze thaw for lysis is that you aren't introducing any buffers or chemicals that could interfere with analysis techniques. For 18S rRNA barcoding one cell is enough, but actually now you could even do whole genome with one cell (even de novo assembly). They don't have cell walls. I don't know much about cilliates, I did research on algae.

@misilagata Год назад

Me encantan vuestros videos. !Encima en español! Mis viajes desde el sofá al microcosmos es una aventura apasionante.

@sajadpp Год назад

Good information 😋

@JeevasJerico13 15 дней назад

This channel is so underrated

@yajuvendrasinghrajpurohit7888 Год назад

James your are amazing and I love this channel

@fatihozdemir6564 Год назад

Awesome Lang translate and Amazing video

@mrwalk6171 Год назад

This would be a perfect organism for single cell whole genome sequencing. Practically what the technology is for, hope this can be revisited one day (i.e. another is found)

@TedToal_TedToal Год назад

Thanks! So, did you sequence its whole genome (and how big is it, btw) or just some part such as ribosomal DNA?

@personzorz Год назад

These days there's no excuse to not get everything

@fasferg2999 Год назад

@@personzorz Using PCR it is much easier, cheaper and faster to just focus upon the 16s Ribosomal DNA than try a single cell full genomic sequencing. The 16S rDNA is typically varied enough that you can determine the taxonomy all the way to family level.

@TheRedKnight101 Год назад

Whole genome sequencing while cheaper than it was just 10 years ago is still fairly expensive. Also you need more than one cell's worth of DNA so you can sequence the genome multiple times to ensure there are no mistakes in the sequencing process. Considering the rarity and size of the organism it's amazing they managed to get enough DNA for PCR. For determining taxanomic classification 18s is usually enough to get a rough understanding of the taxonomy. While whole genome sequencing would provide more information than was needed to answer what it is related too.

@HesderOleh Год назад

@@TheRedKnight101 You can de novo assemble a whole genome from a single cell now a days. It is kind of sad to have that opportunity missed, but honestly unless someone was going to annotate that data, it would be hard to justify the cost.

@travcollier Год назад

@@HesderOleh Whole genome sequencing from single cell is really quite difficult, and even harder to get the long reads or mate-pairs needed to do a decent assembly. There are 'in between' options which amplify a lot of common genes simultaneously instead of just 16s, and then you sequence all those using the same equipment as you'd use for whole genome. That might have been an option here, but it is a lot more expensive. Anyways, this stuff is still quite tricky, even when you have a lot more DNA to start with. FWIW: I do genomics/bioinformatics on mosquitoes.

@janwodarek4133 Год назад

Loved this video, as a future real scientist, present amateur scientist I would love to see more videos about the procedure and method. As I was always drawn to protozoans (now protists i guess) I hope I could one day become another Polish master of microscope. Thanks for the vid :)

@stevenkarnisky411 Год назад

As a non-biologist, I sort of understand the DNA harvesting process, except for the "amplication" process. Is it simply gathering more bits of DNA from the original organism and adding them to the original sample, or does it involve getting the DNA samples to replicate somehow, like bacteria in a petri dish? I wouldn't think the second is possible, but what do I know?

@ptarmigan1356 Год назад

The PCR amplification produces more copies of (certain regions of) the DNA. You mix your ‚template‘ DNA with DNA building blocks and enzymes that can read the template and produce more of it by joining the building blocks together in the correct order.

@clairejeansonne9800 Год назад

Comments for the very, very clean algorithm,

@heartstronggarden5134 Год назад

💕🤘🏼🔬 just chillin'

@cgomes12 Год назад

Congratulations on the amazing job! Which segment of the genome you used in the PCR? Have you considered whole genome amplification and than sequencing of the entire genome?

@timothygreer188 Год назад

Can wait to read the published paper

@hexagonist23 Год назад

So... Where's the DNA of L. Loyezae? Can we download it somewhere? Or is it only available on patreon?

@Bryophytan Год назад

Genbank NCBI

@ooooneeee Год назад

They only did PCR of a part of the ribosome, not a whole genome sequence.

@hexagonist23 Год назад

@@ooooneeee Then they should have titled the video "How We Got The DNA Of A Part of The Ribosome of This Extremely Rare Ciliate"

@finch600 Год назад

I had to clean a mite to mount to be seen under an electron microscope when I was a volunteer. Using a pipette to clean something is so bloody finicky.

@Beryllahawk Год назад

As with the microcosmos, also ourselves. Don't neglect to visit your own "familiar ponds," because in us, too, there is always hidden something more.

@lavinleitrim44 Год назад

The narrator reminds me so much of the one for Tale Foundry, very similar way of speaking.

@husaynbootwala1729 Год назад

If this organism is so rare and has been barely studied, how did you manage to make primers to use in the PCR process? Was the primer a common sequence found in all ciliates?

@sobbski2672 Год назад

I'm assuming they used universal primers targeting the 16S ribosomal subunit

@husaynbootwala1729 Год назад

@@sobbski2672 but would that be enough to get the entire genome? Or would the 16S subunit only be able to amplify some portion of the DNA?

@ptarmigan1356 Год назад

It would amplify a specific region which is quite conserved between organisms. The sequences are similar enough so that you can compare them and see how closely related they are.

@husaynbootwala1729 Год назад

@@ptarmigan1356 but if we wanted to sequence the entire genome, then what would be done?

@mineovernutter Год назад

@@husaynbootwala1729 We would either need to culture the ciliate to get more cells and more DNA. I dont think we are able to culture these cells however otherwise james would have probably done that in this video as a kind of insurance. Or we could maybe do some single cell studies which are quite expensive, and since we dont know which primers are needed to cover the whole genome you would likely not get good enough coverage to get a sequence.

@HNIW Год назад

Gratulejszens myster Dżejms 😁 Pozdro 🇵🇱👍 Powiedz gdzie jest to tajemnicze jeziorko 😉

@mediocreclementine7649 Год назад

I'm going INSANE how tiny is that micropipette?!?

@sharkmeldon Год назад

What a trip. 😁

@pjssjr Год назад

I thought you’d describe a case of horizontal DNA transmission between ciliates and humans!

@RJFerret Год назад

Anyone find online images of "epispathidium papilliferum"? Mine turned up other epispathidium but not those.

@tatoverde9312 Год назад

PCR was abused during the last 2 years. The inventor made it very clear that being + does not mean you are infected or contagious. The use in this experiment make sense. However for diagnosis is faulty to say the least.

@pfischer08 Год назад

Acho que é ótimo ter conteúdo assim em português.

@hherpdderp Год назад

We must avenge the Loyazea

@stonefish1318 Год назад

This was an incredible inside behind microsience! Pls more of this kind

@TheRogueWolf Год назад

I noticed that this video did _not_ feature a microscopic consent form! Scandalous!

@anandg5843 Год назад

@6:15 🙏

@jamesourmasterofmicroscopes Год назад

0:43

@whizthesugoi Год назад

Wooooow

@AlienAnthony Год назад

I have a personal collection of algea I've collected from a few years back. I've lost the original sample from the dirt I got it from but if I wanted to submit a sample could I? I have 3 bottles of it. The first was a sample of the water of the first dirt sample. And each bottle after is from the previous one.

@nightthought2497 Год назад

Calling PCR a pandemic management tool is like calling a hammer a house building tool

@jaikumarsagar Год назад

Bro, what do you use to slow down these microbes? I am facing some difficulty in observing some of these fast swimming microbes under the microscope

@ladycormix4456 Год назад

So why freezing instead of centrifugation? Is the ciliate too small?

@kgv9902 Год назад

It's mind blowing knowing the tentacles are part of the single celled cilliate, i still don't understand how the tentacles move, it ain't got no muscles...

@thea_kober Год назад

I think they're probably moved by the cell's actin filaments or microtubules, like cillia and flagella

@Bryophytan Год назад

@@thea_kober Contractile vacuoles in tandem with cytoskeletal filaments, basically like puppet strings and balloons!

@hallowacko Год назад

Curious: Are there microbes isolated in caves that can be talked about? I started thinking about like, the family tree of eukaryotes, and how something can wind up super rare and only in a couple ponds - like how did they get there? is that the origin of a whole limb of the tree of eukaryotic life? surely not, maybe they just got isolated there or something? Do microbes get isolated? How did eukaryotes get to all these bodies of water?

@DekkarJr Год назад

Ciliate: Can I has dat back pweez :3 :3 ;3

@inkuii Год назад

Out of curiosity, how long did the cycling take?

@sloth0jr Год назад

Yeah, PIA - don't feel comfortable with a VPN service whose marketing slides don't understand the format of an IP address.

@upasananandi1271 Год назад

Ok, so here's something that I've been mulling over for a while: How exactly are the primer sequences for the PCR decided in these cases? As far as I know, the primers not only have to be gene sequence specific, they've also got to be unique to that gene alone and ideally, shouldn't form dimers. That alone creates crazy complications when it comes to designing them. I'm aware that what I know of is sort of the tip of the iceberg, if not entirely wrong, so if someone could offer some clarity, I'd really appreciate it.

@azureabyss538 Год назад

Such a cleanly and evenly posed question, it broke my heart to see it didn't get to see the light. Make sure you pose it in places where it receives appropriate attention.

@Aimee42 Год назад

What do you think of Astrophage?

@kaltkalt2083 Год назад

But what i really wanted to know was how people get the DNA out of one tiny little speck? I figured PCR is used to get a lot more of that DNA.

@Sy-dn9bq Год назад

why exactly did the narration imply the pandemic is past tense?

@CloudColumncat Год назад

Since we are made of cells, it is amazing that we live and are affected by how small and simple cells are. Is it because they have a sense of duty that life is maintained in my body? Is it my will? Maybe my body is like a city. All infrastructure is built using resources by life, and continues to accept, exchange, and survive. In such a daily life, trillions and hundreds of billions of lives arise and die in my body, and new ones are born and maintained. Life is truly a strange thing.

@supreetsahu1964 Год назад

Anyone know the song at 5:50?

@michealwestfall8544 Год назад

RIP.

@Cor97 Год назад

This ciliate measures about 50 um, correct? On my screen it measures 12 cm. So its magnification is 2400, right?

@Bryophytan Год назад

Maths checks out but the ciliate (L. loyezae) is usually 70 um up to 120 um in length

@felvinperez957 Год назад

Actually the answer of this question us very simple. Universal intelligence that if you believe that universe itself is a conscious universe and life on this planet including us human is mere illusion from the conscious universe where we as pary of its spark..

@9a3eedi Год назад

The whole process of moving the organism to a new drop of water sounds like homeopathy :p

@websurfer5772 Год назад

Can you show us proof of the DNA molecule?

@MaryAnnNytowl Год назад

I was sad that such a rare little critter had to die for science. 😭

@belstar1128 Год назад

Rare ? there are more of them than there are humans.

@sdjhgfkshfswdfhskljh3360 Год назад

Is result open source?

@MrBoegela Год назад

i just imagine that some alien takes us up for research and cleans the hell out of us before freezing and thawing and heating and cooling for science soup

@Haplo-san Год назад

I didn't get why freezing and thawing it increases the amount of DNA. It's just a single organism, doesn't it die when you freeze it? And if it dies, how DNA keep replicating itself? I have dozens of questions about it.

@ptarmigan1356 Год назад

Freezing and thawing was just the step to get the DNA accessible by breaking open the cell. It was then amplified by PCR in a separate step to make more copies.

@Quad5Ny Год назад

Audio tracks are still random. They do not default to English/Users Language NOR do they save your preferences, each video When played will pick a RANDOM audio track confusing the viewer!

@alchemi8085 Год назад

Life would be simpler if I were a microbe.

@brianedwards7142 Год назад

You're just being cili.

@nightthought2497 Год назад

Interesting comparison I just noticed. This is a comparison between a real organism and a fictional one. The Legislators in Captive State look kinda smooth in a weird way when "docile" and then look like sea urchins when aggressive. legendrea are like that in reverse. I was thinking about how to turn turn legendrea into a macroscopic alien, only to realize it had kinda already been done.

@liamomalley5779 Год назад

You should be worried about this employee... he puts subjects into isolation, forces them to defecate (probably on camera) transfers them to a clean room and then makes his kill by blowing them apart with ice crystals and cold exposure.... just sayin..