Automation Solutions for Plasmid Construction and Screening

Transcript

0:00	I'm John Fuller, Commercial Product Manager for ECHO Instruments at Beckman Coulter Life Sciences, and Rebecca, would you like to introduce yourself as well?
0:10	Yeah, sure. I'm Rebecca Kreipke. I'm the Field Application Scientist Manager for the Biopharma Portfolio at Molecular Devices, which handles our high
0:23	Perfect. And so, we're doing a little bit of a tag team here, and we're happy to really kick off this discussion on Danaher's solutions for accelerating biotherapeutics discovery, especially that early
0:47	turning that into a product, into a plasmid, automating that, going to a functional assay, and analyzing that, and then taking that cycle again through design, build, test, and learn.
1:05	And so, for the front end of the presentation, we're really focusing on nucleic acid assembly, the applications, the challenges. It's no secret that here at Flagship, there are a number of operating companies that are focused on protein engineering, protein engineering for better antibodies, for better therapeutics.
1:29	Amazing advances that I can't keep up with on non
1:51	The challenge is you can only do so much with a computer, and at some point, you need to start engineering samples. And typically, to feed and to improve those algorithms, you need a large number of samples to build and refine. And this is just an example of some of the combinatorial complexity that you can get when it comes to engineering DNA fragments at scale.
2:20	But we have a solution for that. The solution for being able to economically assemble DNA fragments is with my instrument, the Echo Acoustic Liquid Handler. If you're not familiar with the instrument, what we do is we use sound energy to move a sample that's in a source well.
2:43	So this can be, again, a plasmid in the first well, a DNA fragment in the second well, a second fragment in the third, and so on and so forth. And what we're doing is we're using ultrasonic energy to focus to that source well and dispensing those samples to an inverted assay plate.
3:06	And I'll play the example here. So you put your plate into the instrument. The instrument takes your samples into the plate. There's your assay plate. It gets flipped upside down. I invite you to see one of these in action if you've never seen one live.
3:24	What we're doing first is we're scanning that plate. So you notice that we're not actually making physical contact with your sample. We're pinging a small amount of sound energy into those wells and deriving information about what's in those wells, how thick that fluid is, and using all of this information to come up with a per
3:50	So now that sound focuses and we get, depending on the instrument, a 2.5 or a 25 nanoliter droplet that then gets captured by your inverted assay plate.
4:03	And what this really allows us to do, because the source, the transducer, can move to any position on the plate and the assay plate can move, we can basically generate any well to any well combinations.
4:20	And so this is how our instruments are able to really generate combinations that are just orders of magnitude faster than other typical standard liquid handling instrumentation.
4:34	It's also fairly straightforward to use. So you provide the instrument a CSV pick list that states what well I'm dispensing from, what assay well I'm dispensing to, the transfer volumes, and those are kind of the basics that you need to start that high
4:58	And so what are the things that we can do with this? Well, isothermal assemblies, creating these plasmids, creating mixes and matches of DNA sequences that form a plasmid, and with the Echo 525 Acoustic Liquid Handler, we can take typical assembly volumes and miniaturize those down to as low as 500 nanoliters.
5:24	We've seen that actually go a bit lower, but you eventually just get to a point to where you really don't need to miniaturize any further. It just doesn't make a lot of sense, but you can really get into this nanoliter scale, which not only is cost
5:48	So we've got a plasmid. Rebecca will, in a moment, go into colony picking, but I do think it's important to note that the Echo liquid handler not only does it dispense the DNA fragments itself, the reagents to assemble DNA, but we can actually take transformed bacteria and we can array these with an acoustic liquid handler, with an Echo instrument.
6:15	The image on the bottom is genetically encoded yeast. The image was basically deconvolved into a pick list, and you had, I think, 24,000 spots going onto an SBS Omni tray and dispensed in that pattern, and then they're put in the incubator, and then you get the image.
6:40	A very cool demonstration for the students at the Cold Spring Harbor Synthetic Biology course. This is always a highlight, is taking your college logo or your corporate logo and, you know, making it with bacteria.
6:59	So, speaking of bacterial, I'll turn it over to Rebecca.
7:05	Yes, so thanks for that great kind of front end of the build part of this workflow that we're talking to. So now I'm going to talk sort of through that process of how we at Molecular Devices can help you from either once you get that plate with all of your, you know, colonies arrayed by the Echo or some other form of your transformants, and how we can really help accelerate that plating to colony selection and picking process.
7:31	So, to look at a much more simplified version of this workflow, obviously for, you know, DNA prep or plasmid prep, anything like that, you know, we're pretty familiar, I assume most people, with the steps of this process of assembly transformation, which was nicely covered by John, then moving into your colony plating and picking, and then moving downstream again after that for your, you know, whatever you're doing, your mini prep, your plasmid isolation, any sort of, you know, kind of sequencing or anything like that.
8:01	So, when I talk about automated colony picking, as you can see here, I always like to talk about, we like to start with the problem and not the solution. So, instead of figuring out what the technology is that you want to employ in the lab, really figuring out where in the labs you're having your problem,
8:16	and targeting that with a specific technology solution. And that's why at Molecular Devices, we looked at this part of the workflow, the colony plating to colony picking, and really developed the QPICS, which is our automated high
8:33	So, obviously, video is worth a thousand words. So, anyone who has ever done any sort of manual microbial colony picking, it's a very straightforward, but repetitive and laborious process, where for those of you who have not done this workflow, you're usually taking either toothpicks or some sort of pipette tip, physically touching each colony that's growing on your agar dish,
9:00	and then putting it manually into some sort of liquid growth media. Obviously, this is not something that you necessarily need a whole bunch of training to do, but it is a huge time suck when it comes to what you're having your scientists in the lab spend their time on,
9:18	especially when you're looking at screening tens of thousands of different colonies for whatever your different plasmids or your different gene fragments are that you're looking to screen. So, with the automated colony picker, you're able to basically scale up your throughput ten times. And again, because you're doing this on an automated colony picker, instead of relying on someone who may or may not be fully caffeinated, sitting at the lab bench doing the same thing over and over again, you're also going to reduce your error.
9:47	So, with the QPICS, our picking speed is about 3,000 colonies per hour. So, when paired against sort of your fastest lab techs that are doing this, we estimate their picking is about 300 colonies per hour. And again, then you're introducing the potential of human error into that as well.
10:09	Depending, again, then on how much you're stitching this colony picker together with different pieces of instrumentation, you can either go from as much as 100% hands
10:31	So, for those who have not heard of the QPICS before or aren't familiar with it, it's a very well
10:47	So, it is the fastest colony picker on the market, as I said, about 3,000 colonies per hour. And the nice thing about this being in our hands and really being, you know, a molecular device is we think of ourselves as a biology company, as well as a technology company.
11:02	So, we're able to keep the scientific expertise in terms of continually pumping that into the development of this instrument to be able to continue to meet the needs of this workflow as it changes over time.
11:14	So, just a little bit about what is in the QPICS. The brains of the operation is this actuator head. This is where you're going to have a high
11:33	We also have organism
11:55	You also have your source plates. So, this is where we can have any number of different formats of plates from your Q
12:09	It does come equipped with a barcode reader, so you can always track all of your data, as well as an in
12:27	So, especially if you have any sort of 2025 environmental pledges that might be coming up, this is a good place to start looking to decrease some of that waste.
12:39	So, the way it works, you take your agar trays, you put them into the QPICS, it's going to image your colonies. The software in the QPICS will analyze those colonies based on your picking parameters.
12:52	It will pick all of the colonies that you tell it to, inoculate into your liquid growth medium, sterilize the pins, and then just continue through that cycle until all of your pins or all of your colonies have been picked.
13:03	It's really based around this very easy application
13:15	When we train people, usually you're up and running within a few hours, and certainly, you know, depending on the level of complication in your workflow, within a day, usually.
13:27	Again, it's all image
13:46	The software will then automatically analyze which colonies meet your parameters to go ahead and pick those.
13:52	It does also come with the option to do fluorescence. So, if you have any sort of like GFP indicator that you want to pick, you know, high levels of GFP or low levels of GFP, you can also specify that within the software.
14:04	As I said, in addition to the plating or the picking, we do have several other applications. So, we can also equip it to do your plating.
14:18	So, if you want to do something at a little more high volume than what John was speaking to with the Echo, we can put a single
14:31	We then have, again, purpose
14:48	We also have the ability to do lots of different library management. So, in terms of if you are going from a 384
15:14	And the nice thing about this, again, so you can have that standalone by itself if you just want to get the, you know, quickest up and running where you're having a manual operator interact with this, but where, in my opinion, it gets really fun is where we start integrating this with other pieces of instrumentation, as I mentioned.
15:33	So, you can pull the back off of this instrument, and now it's going to allow a robotic arm to feed all of your plates onto the deck of the QPICS bed and really be able to use this as sort of the start of building a total walkaway work cell.
15:49	So, we can start really basic. This is an example of some of our most basic QPICS system. So, this is just where you have some ambient hotels. It's basically taking the place of an operator who's feeding those plates in from the front where you're doing, you know, you can see I diagrammed out sort of this workflow here at the bottom from assembly, transformation, plating, growing, picking, growing, and then downstream to your, you know, let's say mini prep.
16:13	So, the places in green are where you're automating, and then everything else is going to be offline. And then what's nice is we can work with you on your workflow to sort of stack this together with additional instrumentation and sort of build up with as many other, you know, other pieces of instrumentation to really cover as much or as little of that sort of plating to picking workflow, as well as with your incubation as you need.
16:39	And then as, you know, Greg had set up in his introduction, as well as John, where the power really then can be derived is when we start partnering with our sister APCOs within Danaher to be able to really build either, you know, one entire workflow or we have some different, you know, islands of automation that we've done where you look at different pieces of the workflow and go from really just one part of it to being able to give you a total end
17:09	And so, then to sort of pick up again from where I have left off at the end of our colony picking, I'm going to hand it back to John again to talk about the downstream of that applications back at Beckman.
17:25	Thank you, Rebecca.
17:27	So, now with the QPICS, we've engineered, we've picked a lot of colonies, we have a lot of plasmid prep to do.
17:35	Oh, sorry.
17:38	And so, an offering that we have at Beckman Coulter Life Sciences this year that I'd like to focus on, kind of with that concept that Rebecca mentioned with workflow automation, islands of automation, we have this, which is the Biomek ECHO1 high
17:59	But this enables, so I mentioned the ECHO Acoustic Liquid Handler using that non
18:14	ECHO instruments are amazing, but they can't do everything in a liquid handling workflow. And this is why we have it paired with a Biomek I7 hybrid workstation.
18:24	So, the components on the left as part of this package are included, the ECHO instrument, the ECHO software.
18:32	We have a Biomek I7, which features two liquid handling, primary liquid handling dispense options, a 384
18:47	And then on the right, we have a SPAN
19:02	So, one thing, though, you look at a system like this, and if you're not an automation engineer, some people might get a little bit daunted by this.
19:11	And we don't want them to because what we want to be able to show is we have really sensible entry points into automation.
19:21	You know, we can start with a simple stand
19:41	So, when I was in field applications for LabSight, one of my favorite applications to really get someone familiar with the power of the ECHO liquid handler was a qPCR reaction setup.
19:55	I could typically take a customer in about a half an hour, show them a reaction, and then have them do a qPCR reaction.
20:00	My last thing, if you'd like to see some very significant developments to SDM,
20:05	this again is very simple to tell where to look.
20:09	So, I want to say thank you to the facilitators,
20:12	and I look forward to working with you for the qPCR
20:16	ongoing hybrid solutions.
20:18	Thanks.
20:21	Thank you, David.
20:24	Thanks, Cole.
20:24	And Joe.
20:25	So, we have a standalone application.
20:28	We can expand that application to increase throughput
20:31	by integrating the Echo and the Biomek instruments together,
20:35	and then really scale up and expand the workflow to where you're headed.
20:42	So, when I was in field applications for LabSight,
20:46	one of my favorite applications to really get someone familiar
20:51	with the power of the ECHO liquid handler was a qPCR reaction.
20:56	I could typically take a customer in about a half an hour,
20:59	show them how the ECHO instrument works
21:04	with the ECHO plate reformat software,
21:07	show them that they can simply use the graphical interface
21:11	to lay out where their primers are arrayed,
21:17	where their cDNA samples are arrayed,
21:19	and set up, again, in a matter of minutes, a simple PCR workflow.
21:26	Customers do that.
21:28	You feed the instrument the plates.
21:31	You get fully assembled PCR reactions coming out.
21:34	You seal those up, and then you start to see amazing data
21:39	coming at scale and miniaturized.
21:42	And what happens is people tend to get jealous of their time,
21:45	and they have better things to do than loading the ECHO PCR plates.
21:53	And so that's where this comes in.
21:55	We can take that standalone application,
21:57	and we can slide and rotate the ECHO instrument
22:01	to integrate with the Biomek instrument.
22:06	So, in the next phase, what we can do
22:09	is we can take a straightforward Biomek method,
22:13	and if you're not familiar with the Biomek liquid handling software,
22:19	of the tip
22:22	it's fairly straightforward for creating these methods,
22:25	and you can create a method where you have a deck layout
22:30	where all the samples are located, and you can tell that Biomek,
22:35	this is the ECHO PCR method that we're going to run today,
22:41	and the Biomek will feed the ECHO the plates,
22:45	do any insert swaps, any centrifuging that needs to happen,
22:51	and so now you can really start to run these applications at scale.
22:59	The real power is bringing everything together for the DNA,
23:04	the plasmid, the mini preps,
23:07	as well as high
23:12	and really, the power of the Biomek instrument,
23:16	it's not just the automation,
23:18	but it's really this software that allows you to prevent user errors.
23:24	We have a couple of ways that this happens.
23:30	The one on the left is guided labware setup
23:33	where it actually will walk your end user through
23:35	to ensure all of the labware is set up for the method correctly,
23:41	and then deck optics final check actually uses cameras on board the Biomek
23:46	to ensure that you've put those correct plates,
23:50	those correct tip boxes in the correct locations,
23:53	and hopefully can prevent an execution problem before the run happens.
24:00	Then of course, then we're in walkaway mode
24:03	where you're really able to run these multi
24:07	and we've got remote monitoring along the way.
24:11	I think Rebecca touched on this.
24:15	One of the issues that we get when we consult with customers,
24:20	we talk about throughput, we talk about your needs,
24:22	we ask, well, how many samples do you plan on running in a day?
24:29	The answer can be, well, honestly, we don't know.
24:32	We don't know where we're going to be in a year.
24:33	There's just so many variables.
24:36	What we typically say is, ideally, it's really good to know
24:40	what that throughput looks like so we can build the solution now,
24:44	but we can also work with you to expand it as the needs become more apparent.
24:49	As the needs shift, we can change the solution
24:54	to fit the throughput and the needs at the time.
24:59	Again, this particular Biomek ECHO1 offering,
25:02	which I'll invite you to actually come see a miniature model here
25:09	that's outside on the show floor,
25:16	and you can actually see what this looks like in person.
25:21	I'm going to finish the presentation.
25:23	We talked about DNA engineering.
25:26	We've talked about colony picking, plasmid assembly, sequencing.
25:30	Well, what are some of the things that we can do with DNA?
25:34	And that's a technology that I think is evolving rapidly,
25:39	and that's cell
25:41	The cell
25:44	typically, if you're doing mammalian engineering,
25:47	the design, build, test, learn cycle can take weeks.
25:50	If you're needing to get that plasmid into a mammalian cell,
25:54	you need to select that cell, you need to culture the cell,
25:58	and then you start getting that protein product.
26:01	It takes a lot of time.
26:03	One option that you have is actually doing protein synthesis in vitro
26:09	with cell
26:12	We don't just collaborate, obviously, with our Danaher sister OpCos.
26:17	Had a nice collaboration with New England Biolabs
26:20	on miniaturizing NEB Express and Pure Express cell
26:26	getting those miniaturized down to as low as 500 nanoliters
26:30	to automate that protein expression.
26:33	And you can take that DNA,
26:35	and you do a number of things in a number of hours.
26:41	And this is just a quick customer showcase
26:44	talking about using automated miniaturization
26:49	of cell
26:53	that a customer of ours published not long ago,
26:57	presented on a webinar.
26:59	So they were using minibinders for high
27:06	for COVID spike proteins.
27:08	And the takeaway here was, you know,
27:10	a standard antibody expression workflow takes weeks.
27:13	You can get this down to a number of hours to days,
27:17	miniaturize the cell
27:21	down to two microliters,
27:22	and really expand and permute the design space with cell
27:28	You could then take those cell
27:30	and we'll talk about it here in this topic,
27:33	but, you know, pass that along down to the Molecular Devices
27:37	plate reader for plate
27:40	or functional screening with an Echo
27:45	We will stop there for today and happy to take any questions.