| 00:09 | Oh, yeah. Ok, welcome. Um Sorry for the, |
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| 00:31 | uh delay. I left my office back way ended up in this prison |
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| 00:37 | . If you, if you don't there's a big fence put around that |
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| 00:41 | , what it's called. And I in there and I go, how |
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| 00:43 | I get out? And so I'd walk around and finally found an |
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| 00:47 | So anyway, uh, so beware you get trapped in there. |
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| 00:54 | let's see. Ok. Uh if, um, if you're one |
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| 01:01 | took the exam at the dart All right. So they, |
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| 01:06 | they just sent me the exams this . So those will be graded today |
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| 01:11 | posted them on the canvas, If you've been waiting for wondering what's |
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| 01:16 | on there, that's what's going Ok. So, uh, let's |
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| 01:21 | . So usual stuff. Uh, , weekly quiz Friday through Monday. |
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| 01:29 | work. Uh, four is divided two parts. The part one, |
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| 01:34 | , we'll complete that today. Um two is kind of next week. |
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| 01:41 | , uh, let's see. So break coming up. Uh, so |
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| 01:44 | got next week and then break for week. Um So getting the |
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| 01:51 | The exam is the, I forget the, I think it's, |
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| 01:59 | we come back from spring break then think the exams a week after |
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| 02:02 | I think, uh, but I'll check on that and, and |
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| 02:07 | it in the email coming up So, uh, which means for |
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| 02:12 | that the scheduling thing you want to aware of when that opens? So |
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| 02:15 | check on that this afternoon. I that's everything. Um Any questions about |
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| 02:24 | ? Yeah. Yeah. OK. chapter four. So again, kind |
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| 02:32 | it's probably worth spending a minute Um Let me turn this down a |
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| 02:37 | here. Uh OK. So right? Microbial growth, bacterial |
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| 02:45 | Um We looked at, so I is kind of a, a road |
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| 02:49 | of what we're doing here. So you're gonna grow microbes, so what |
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| 02:53 | they need to grow? We already about the I call it chomps. |
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| 02:58 | that's, that's what the M cho chomp. It's hard to say chomps |
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| 03:04 | an N in the middle. But , um that's uh what we gotta |
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| 03:10 | if we're gonna grow these things, ? So, um and you divide |
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| 03:15 | up into those that you need lots macro and just little amounts of micro |
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| 03:20 | like trace elements, Cobalt, stuff like that, uh certain |
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| 03:26 | Um But certainly the big influence here gonna be carbon in terms of how |
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| 03:31 | yield you get yield being how many I'm ending up with at the |
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| 03:36 | OK. Um And so uh the nutritional types, right? Uh or |
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| 03:43 | one a photo a Ditro what have that obviously changes what you're gonna do |
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| 03:50 | terms of ingredients. Um And then form, right? Are you gonna |
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| 03:57 | it as a solid liquid, Plates or liquid culture? So there's |
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| 04:02 | for that. Um the type we'll about that in a second here. |
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| 04:07 | versus defined. OK. And then can, um and there's reasons to |
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| 04:13 | things like uh selective media differential, you did if you're in lab did |
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| 04:18 | last week. Web six. Um also things like enrichment and media for |
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| 04:24 | uses. OK. So just to let's see, is there anything |
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| 04:29 | So let's uh just a second on , why it grows stuff well, |
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| 04:35 | wide microbes. Uh Well, obviously you're a ecology person studying birds or |
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| 04:42 | , large animals, that's of no to you. Although um you will |
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| 04:47 | know certainly nutrients, um whether you're or us, you know, certain |
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| 04:57 | um are of course critical right to health and many other things. So |
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| 05:03 | definitely has an impact if one is ecology and looking and seeing what's out |
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| 05:07 | and studying populations and things. How our food sources impacting those populations, |
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| 05:13 | like that, right. But we're, we're not care, we |
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| 05:16 | care about those things we can see our naked eye, right? We're |
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| 05:19 | at microbes here, right? uh so why, so why do |
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| 05:22 | grow these? Well, if you're of the biotech people, right, |
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| 05:27 | be working on a job where you'll growing this probably at large scale, |
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| 05:32 | ? So if you are one that um I mean, even a small |
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| 05:36 | in the academic lab, right? grow stuff to, typically to get |
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| 05:41 | and do stuff with DNA. They're it for sequencing a gene you're looking |
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| 05:45 | or maybe it's for taxonomic purposes, need to isolate DNA. That's oftentimes |
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| 05:51 | the white cells are grown lab for purpose or for some particular protein and |
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| 05:55 | enzymes. Ok? Um Now, know, in the lab scale, |
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| 06:02 | ? You're only doing small quantities of you don't need that much, |
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| 06:05 | So most molecular biologists um don't really about getting super high cell use, |
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| 06:12 | ? That's why they use things like ar, this thing called lb, |
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| 06:16 | is very similar, right? It's the they have these kind of what |
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| 06:19 | call complex nutrients, which are basically of an all in one thing, |
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| 06:23 | ? If you give a protein as nutrient, you know, you're gonna |
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| 06:28 | cho nps in that one ingredient, ? So it's kind of covers all |
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| 06:35 | bases. Ok? Generally speaking, , just proteins by themselves are not |
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| 06:41 | great way to get lots of right? It has to do with |
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| 06:44 | you want carbon, you want more in there, right? Protein provides |
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| 06:48 | , but not in the same Ok. So, aside from |
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| 06:53 | so that's why most, most that this kind of, uh, I |
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| 06:56 | , I'm just throwing it for, see some DNA or some protein. |
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| 07:00 | all they do. They want to a lot of time on that |
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| 07:02 | But they know it supplies this and grow and they'll just grow it |
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| 07:07 | right? If you're in industry, want to be more quantitative about |
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| 07:11 | right? More meticulous about it. so because your goal there is to |
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| 07:15 | lots of material because lots of cells lots of your product, right? |
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| 07:21 | it protein, what have you. then you are interested in, |
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| 07:25 | you know, being more detailed about if you will, right? So |
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| 07:29 | where you know, you dissect your , right? Providing the things that |
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| 07:35 | know, we can readily use and fast on, right? And you |
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| 07:38 | kind of preliminary experience experiments to figure out. But again, you're, |
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| 07:43 | focused on um um how can I these things to really grow fast and |
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| 07:48 | my product? Right. And it be that you have a a base |
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| 07:54 | that can grow yourselves well, but you need to add something in there |
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| 07:58 | promote the specific formation of the particular that can sometimes happen to you. |
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| 08:05 | you may grow them up the high and you go OK, I'm gonna |
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| 08:08 | X now because I know that will lots of production in our protein, |
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| 08:12 | stuff like that, right? You be working with strains that are engineered |
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| 08:17 | different ways. When you're in you're looking at um production of |
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| 08:21 | what thats to make a particular you want to make a ton of |
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| 08:25 | , right? So not only do use the ability to manipulate growth but |
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| 08:32 | um you manipulate the genes, So we'll learn about this stuff next |
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| 08:38 | . But um uh say gene X a protein you want or there are |
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| 08:43 | you can do at the DNA right? You can influence expression of |
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| 08:48 | gene and ramp that up. So do that plus lots of cells |
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| 08:54 | that's gets you to your goal. that's often what you do in industry |
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| 09:00 | do both the growth thing and then do, let's manipulate genes and I've |
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| 09:04 | both of those things in industry. they all go together. OK. |
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| 09:10 | , um but for academic lab, just kind of, I just need |
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| 09:13 | CS to get, you know, all that effort into it. |
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| 09:17 | Typically, OK. Anyway, so kind of uh you know, why |
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| 09:21 | might grow stuff, OK. As . The uh we're talking about this |
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| 09:29 | uh at the end today, next time uh cell numbers grow, |
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| 09:33 | talk about cell numbers. OK? growth rate grows curve. It's very |
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| 09:37 | to get a P, what's the of growth of this thing? |
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| 09:42 | Because uh I need to make sure when I'm scaling this thing up, |
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| 09:47 | , scaling up can be like from big to like as volume that's as |
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| 09:54 | as that, you know, 100,000 in some cases. OK. So |
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| 10:00 | gotta know, you don't want to experiments like that at 100,000 gallon |
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| 10:04 | OK? You wanna do it at small scale and then translate it |
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| 10:08 | transition up. OK? Because there's be stuff's gonna happen, right? |
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| 10:14 | , you, you need to figure out at a small scale. Not |
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| 10:17 | big because remember the industry, everything money, right? If you're doing |
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| 10:23 | super big, that's lots of money . So do your experiments at this |
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| 10:28 | and then transition up, right? um liquid solid, right? And |
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| 10:34 | course, when you're trying to do of cells, it's gonna be a |
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| 10:38 | because you can control the volumes. right, it's more practical to do |
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| 10:41 | in liquid obviously on the plate. . So plate has its role in |
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| 10:48 | the pure culture, maintaining the pure . That's where, that's where that |
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| 10:53 | is at. OK. So different forms for different purposes. OK. |
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| 11:00 | All right. So let's look at um stupid here. OK. So |
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| 11:11 | talk about this. So again, factors, nutritional types. So remember |
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| 11:16 | the you can, you can supply essential nutrients. Ok. That may |
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| 11:21 | enough for most things to grow and happy, right? But there's typically |
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| 11:25 | be some that if you are deficient certain pathways, they need some additional |
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| 11:31 | if you will. So you can that in the form of vitamins that |
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| 11:34 | they can't make um amino acids they make, but in the medium for |
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| 11:40 | , uh it's like us, we , we can't synthesize all our |
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| 11:44 | our amino acids, we have to those in our diet. So very |
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| 11:47 | to that. Uh blood and serum very common additions for growing pathogens. |
|
| 11:54 | ? Uh They u they use these from your body, you know, |
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| 11:58 | we don't necessarily maybe know what the thing in the blood that makes it |
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| 12:02 | or in the serum because both of things contain lots of chemicals. And |
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| 12:08 | you say I don't care about but I know adding blood helps. |
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| 12:12 | that's what you do. OK. OK. Oxo. Remember that Oxo |
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| 12:17 | the one that's deficient, right? you'll simply say, OK, |
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| 12:21 | this, this bug is a histidine troph, meaning it can't make that |
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| 12:27 | acid, right? So this word is always used in the context of |
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| 12:33 | blank Oxo and whatever the pathway is and that's what you fill in the |
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| 12:38 | with you. Uh All right, talked about this at the end last |
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| 12:44 | . So I think we're probably pretty with this and some of these terms |
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| 12:48 | overlap chemo organo trope or simply organo , chemo heterotrophic, same thing, |
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| 12:55 | , chemo Autotroph, same thing. Just saying litho it is pretty much |
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| 13:01 | , it's the same thing. Uh are both different, right? They |
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| 13:05 | use light of course, but they in terms of the carbon usage, |
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| 13:09 | ? So like hetero hetero auto Energy source, light, chemical oxidation |
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| 13:16 | source. So again, overlap here then uh so remember the heterotrophic units |
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| 13:27 | make stuff with plus getting energy from . OK. So um so I'm |
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| 13:36 | to dive into different types of growth . OK. And most all these |
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| 13:43 | are just stuff that many of these just like an all in one powder |
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| 13:47 | you just boil out and dump it water and boil it and auto play |
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| 13:51 | and dispense it. Others may be uh laborious to make where you have |
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| 13:58 | add this chemical, this chemical, chemical, this chemical volume so |
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| 14:02 | So it it can vary depending on you're making. OK. So one |
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| 14:08 | , so here we're gonna do some calculations a bit. Nothing complicated. |
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| 14:16 | . Nothing beyond subtraction, division, , multiplication and log the base |
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| 14:23 | OK. Um But you will, will be like two or three of |
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| 14:27 | on the exam. You'll see them a, in a quiz as |
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| 14:31 | Um There's practice problems on canvas as that are all worked out. Um |
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| 14:37 | will go through some of these couple these today, but you'll have, |
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| 14:41 | have be able to take a calculator you to, to testing center and |
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| 14:45 | it. I've done it every semester I make them aware of it. |
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| 14:49 | , beforehand, so you don't get about it. Uh So just know |
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| 14:53 | and I don't care what kind of it is. That's so that's, |
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| 14:57 | I'll mention this again as the exam up. So just know that. |
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| 15:01 | . So let's go on to growth . OK. So let's look at |
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| 15:04 | question here. OK. So what of gross medium is this? |
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| 15:12 | Um Define complex selective or differential. . Here are all your ingredients. |
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| 15:28 | Oops, sorry. Let me tell . OK. Cutting down here. |
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| 16:01 | 543 due to dramatic pause. Come . OK. Uh BB is the |
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| 16:19 | B is correct. So what, in this box makes the complex |
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| 16:28 | right? The the peptone, the extract, right? So if we |
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| 16:34 | those out, OK, we just these altogether. Um that and that |
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| 16:44 | this would be a defined medium, ? That would be a defined Xing |
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| 16:51 | those two things. OK? Because could, I could put a periodic |
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| 16:55 | there and give you a calculator and could figure out the exact moles of |
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| 17:00 | atom in the medium, right? the, that's the nature of a |
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| 17:05 | medium, you can define every single and its quantity in the medium, |
|
| 17:10 | ? It's what you use. If gonna do a growth study and go |
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| 17:15 | ? What are the gross requirements of thing I'm growing? That's what you |
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| 17:19 | do is use something like this because wanna control what every little atom in |
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| 17:23 | is and how much you got. . So now the complex medium as |
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| 17:30 | uh is one that has one or of these. OK. So it |
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| 17:37 | matter that you got a bunch of constituents that you might go. |
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| 17:41 | That's defined no, as soon as put even one of these in |
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| 17:46 | It becomes complex. OK? Because you know that, so basically |
|
| 17:52 | beef extract, um soy based products plants are used in the same |
|
| 17:59 | Um what are called infusions, there's heart infusion. Uh these are all |
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| 18:07 | not to be crude, but they come from the slaughterhouse, right? |
|
| 18:11 | so all the stuff left over, ? Which are basically meat products of |
|
| 18:16 | types, more from plants, plant , they basically either boil them, |
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| 18:22 | they um add enzymes to them to them down further. Uh But collectively |
|
| 18:29 | really just rich in protein, Rich in protein. And so you |
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| 18:33 | that by providing these things, you're provide, you know, cho NPS |
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| 18:39 | lots of preformed stuff, right? amino acids like um vitamins, et |
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| 18:47 | , right? Are gonna be in . Right. So, yeah, |
|
| 18:51 | of this body if I took this and chopped it up and boiled |
|
| 18:55 | All right. And added it to gross medium. What's in here? |
|
| 18:58 | right. All those things, amino acids, right? Fat, |
|
| 19:03 | . So it all goes in, all part of it. Right. |
|
| 19:07 | , uh, but you don't right. What you don't know |
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| 19:11 | you know that all that stuff is there. You just don't know the |
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| 19:14 | down to the grain per liter quality what's in there. OK. You |
|
| 19:19 | , these things are in there but the exact quantities like you do in |
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| 19:23 | def defined me. Right? that's the difference between those two |
|
| 19:28 | Um So, uh here is just of the way. So here's just |
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| 19:34 | three recipes. OK? Um So uh lb that's that one that the |
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| 19:44 | biologists use. It's very easy. , it has uh uh nothing but |
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| 19:49 | , these complex ingredients. Crypton used . Um And that all these, |
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| 19:55 | we call powdered media, they, rehydrate them according to directions and they |
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| 20:00 | to the right. Ph. So you don't have to worry about |
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| 20:04 | or solu concentration all goes, it's calibrated to go to the right |
|
| 20:09 | OK? Um Because you don't even , you don't even have to use |
|
| 20:13 | Ph. Here, it's all just right to the right thing. |
|
| 20:16 | So, um, so completely complex here. Uh the M nine so |
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| 20:24 | , you can see every constituent, course, glucose is H six C |
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| 20:28 | H 1206, right? So uh know everything in there and the qualities |
|
| 20:34 | ? Uh Similarly this is the five , OK? But note the difference |
|
| 20:40 | these two, OK? When you're to go, OK. What could |
|
| 20:45 | on this, right? What could on this medium are these media you |
|
| 20:50 | at always focus on this first see , OK. And so lb these |
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| 21:01 | complex organic forms, right? That's I you could grow on this. |
|
| 21:08 | ? I wouldn't taste very good but could grow on it. Um M |
|
| 21:13 | . OK. C source is glucose . That's again, heterotrophic trope, |
|
| 21:20 | ? They could grow on uh these two but then you go down to |
|
| 21:25 | , right? So for oxidizers, remember they use inorganic compounds for |
|
| 21:31 | OK. And so they can use like elemental sulfur. This is their |
|
| 21:39 | . Uh but the carbon is this ? CO2 or you might see it |
|
| 21:47 | uh co three a carbonate form. . So CO2 of course, is |
|
| 21:55 | , you can bubble it in uh it's on the plate, you can |
|
| 21:58 | kind of be it's in the right? Um But you can also |
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| 22:03 | it as a carbonate uh form. very often cells that are autotrophs that |
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| 22:08 | CO2. Uh they can take CO and they have an enzyme that converts |
|
| 22:13 | to CO2 that they use to OK. So if you see that |
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| 22:19 | in medium, uh of course, is obvious if you see CO2 |
|
| 22:24 | right? But if you see something this H two co three, that |
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| 22:29 | is, is gonna be a CO2 , OK. Uh nutrient auger, |
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| 22:34 | you use all the time in right? Peptone def extra tract |
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| 22:37 | Completely. Again. Um now heterotrophic that you know the complex nutrients |
|
| 22:44 | complex organic nutrients. OK. Um Let's see. OK. So |
|
| 22:52 | we call complex medium or rich OK. Rich because because of these |
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| 23:00 | nutrients, they provide lots of preform . So remember, right, if |
|
| 23:04 | just had to think in terms of nine, OK. M nine versus |
|
| 23:12 | . OK. What which, which one would the cells grow faster |
|
| 23:20 | more than likely? Maybe I achieved same here but grow like this versus |
|
| 23:29 | the lb or the minimum medium? one? Yeah, lb because it's |
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| 23:37 | preform stuffs that here. I don't to synthesize this. It's already made |
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| 23:41 | me. Whenever a cell has to genes turn on pathways in order to |
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| 23:47 | something that's time and energy. If it's already getting preformed stuff to |
|
| 23:54 | , it doesn't have to synthesize anything time it can grow. So there's |
|
| 23:59 | a time difference between growth on minimum and complex. OK. Um Now |
|
| 24:08 | might achieve ultimately a higher final cell on this one. OK? Depends |
|
| 24:19 | how much carbon you're adding. So that's the big influencer what you |
|
| 24:24 | get, whether you're growing like this or that rate or this rate where |
|
| 24:30 | end up is, is all based how much carbon are you adding? |
|
| 24:35 | . That's ultimately the determiner of uh you're a complex or millimeter that plateau |
|
| 24:43 | you reach is this carbon that tells how much you're gonna get there. |
|
| 24:48 | ? Um And you can do if so inclined, you can do experiments |
|
| 24:53 | OK. How much you know in 2 g, 100 glucose, how |
|
| 24:57 | cells that actually get me? You quantitate that stuff, it's easy to |
|
| 25:01 | , right? So these are things keep, you keep note of if |
|
| 25:05 | going to scale up, right? gonna predict I wanna get this many |
|
| 25:10 | , then you know how much carbon very simple, right? Stuff you |
|
| 25:14 | stuff you do all the time in . Um OK. So from a |
|
| 25:20 | standpoint though, what's done? the the the strict defined medium has |
|
| 25:27 | role in in different uh experimental OK. If you want to know |
|
| 25:33 | what the nutrient requirements are, you have something like this because you control |
|
| 25:37 | and you know exactly what amounts of atom. OK. If you are |
|
| 25:44 | also use middle medium in um in experiments or maybe you're looking for um |
|
| 25:52 | pathway, uh identifying uh parts of metabolic pathway or something. You're also |
|
| 25:59 | need to control that. Right. so you want to know exactly what |
|
| 26:04 | eating to affect expression and what So, there are contexts where you |
|
| 26:09 | have minimum meaning for that. If you are one that is, |
|
| 26:17 | , and that is, uh your is I wanna get lots of |
|
| 26:20 | then what you're gonna do is a really of, well, it's complex |
|
| 26:25 | . You're gonna use a complex but you're gonna use one that's something |
|
| 26:28 | M nine, right? And you're crank up the glucose levels amounts and |
|
| 26:35 | gonna add a peptone or a one these things because you know, that |
|
| 26:39 | help you boost, boost growth in of giving it, you know, |
|
| 26:42 | and vitamins and things. But you're that glucose that carbon to raise up |
|
| 26:49 | cell density. Ok? In the days they used to call it semi |
|
| 26:55 | . OK? Now we just call complex. OK? Uh But your |
|
| 26:59 | medium, the one in the middle , we also call it synthetic. |
|
| 27:06 | which I'm not sure why, but medium often used. So you know |
|
| 27:10 | there's gonna be gross differences because on minimum medium, it's minimal because they |
|
| 27:17 | to the cells growing and they have make all their stuff from those base |
|
| 27:23 | , right? Um They have to their vitamins, they are amino |
|
| 27:27 | right? So that, that growth , that growth rate is gonna be |
|
| 27:30 | little bit slower because of that. . On a complex medium, they're |
|
| 27:35 | these things and so they can grow rate. OK? But the |
|
| 27:40 | the ultimate final yield amount of salts determiner is the amount of carbon in |
|
| 27:49 | medium. OK. That makes OK. So we do the biotech |
|
| 27:56 | write this stuff down because you're not get in a textbook. Ok. |
|
| 28:02 | , let's see. Ok. Does anybody know what the word fastidious |
|
| 28:08 | ? Bye. You ever have an friend or friends that, um, |
|
| 28:17 | , you're at a restaurant and they a salad with chicken on top? |
|
| 28:24 | . They say they tell the uh, where was the chicken |
|
| 28:28 | Was it like, uh, free ? Uh, where at exactly? |
|
| 28:32 | . Uh, I don't want um, tomatoes on my salad. |
|
| 28:36 | don't want this. And do you this, that they're very exacting? |
|
| 28:40 | high maintenance? Right. Those are fastidious people, right? So, |
|
| 28:46 | time if your friends, one of , tell them you're so fastidious and |
|
| 28:49 | what they say. Ok. so similar with bacteria that they are |
|
| 28:55 | , those that aren't, are pretty to grow and maintain. Those are |
|
| 29:00 | , I'll come back to that And this one, ok. In |
|
| 29:04 | one, I'll come back to that too. This is fastidious. |
|
| 29:09 | See, see, this is the stuff we used to see, but |
|
| 29:12 | look at all this stuff, When you're making a media for fastidious |
|
| 29:16 | , you're gonna be in the lab day because you're gonna weigh out this |
|
| 29:18 | this and this and that and the and this and that there's a lot |
|
| 29:21 | requirements to grow, ok? Which very annoying if you have to work |
|
| 29:26 | these types, right? So um the nature of invest has, has |
|
| 29:30 | lot of requirements in order for it grow and you can see the, |
|
| 29:34 | whole list of stuff over here, ? So OK, let's go |
|
| 29:41 | OK. So this question, so gonna do some reading here. |
|
| 29:47 | So each of the following ABC D represents the chemical composition of various growths |
|
| 29:56 | . OK? Which one is a defined uh minimal medium suitable for growth |
|
| 30:04 | a heterotrophic Procar? OK. And let's uh forget about E hm. |
|
| 30:24 | . So you're looking for chemically defined minimum medium. OK. Growing ahead |
|
| 30:29 | trope. OK. There's some kind warning going on. That's like one |
|
| 30:36 | those emergency things. No, I get it. OK. Oh, |
|
| 30:47 | would help if I do that. sorry. Oops, goodness. Very |
|
| 30:54 | screen here. There we go. . OK. So defined mela medium |
|
| 31:19 | trope. OK. OK. Count 765 43 and there's such a delay |
|
| 31:52 | . I don't know why you get . Ok. Well, I'd say |
|
| 31:59 | and disappears. Goodness, stop OK. All right. I screwed |
|
| 32:08 | up. Uh So all together. , the correct answer is which |
|
| 32:15 | B Yeah. B it's B So we can eliminate uh DD right |
|
| 32:21 | the bat, right? Because I you the hint here of carbonates or |
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| 32:25 | atmospheric sources of CO2. So that's an autotroph, right? So |
|
| 32:29 | eliminates D, so we got A and C, so we see A |
|
| 32:35 | C have these complex nutrients in These extract beef extract, uh A |
|
| 32:42 | a soy digest, right? So just one, right? It has |
|
| 32:46 | one of those complex nutrients but that makes it a complex medium. |
|
| 32:52 | . So this and uh these, ? So this is the only one |
|
| 32:59 | completely a defined medium. OK. , and can grow a hetero, |
|
| 33:04 | ? So the, the sucrose, glucose or fructose all complex organic |
|
| 33:09 | That's what a hetero would like. B is the only one that's |
|
| 33:12 | OK. Um Let's look at this . OK. So OK. So |
|
| 33:20 | the oxo troph definition. So would bacterium known to be a histidine oxo |
|
| 33:27 | able to grow on this medium? or no. OK. Speed this |
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| 34:09 | a little bit here. All Counting down from 10. Yeah. |
|
| 34:25 | I popped up. OK. So see. So who, who |
|
| 34:32 | Yes. OK. Why is Yes. You know why it's |
|
| 34:42 | Why is it? Yes. So what does that mean? So |
|
| 34:51 | what's can, can it or can not make histadine cannot write? |
|
| 35:00 | uh, the ox is, you that you go, OK. It's |
|
| 35:04 | in something what's deficient in, it's the term is. Um, |
|
| 35:08 | So here's the nox atrop A B , vitamin oxy troph or whatever |
|
| 35:14 | that's gonna tell you deficient. can't do it. So, |
|
| 35:19 | who else answered? Yes. So, so why is it? |
|
| 35:24 | . Yes. Yeah. exactly. . So it's the, the complex |
|
| 35:30 | you could pretty well assume will be to provide the history. Ok. |
|
| 35:35 | remember these complex nutrients, if if you, if you see the |
|
| 35:39 | just peptone effects and these things just of, think of yourself, |
|
| 35:45 | And you are the ingredient now going the pot, right? Histadine is |
|
| 35:50 | be in here somewhere, right? are other vitamins and other things, |
|
| 35:56 | ? So that's gonna supply the, . So if you were trying to |
|
| 36:00 | a histadine oxo, right? And the nature of that or whatever, |
|
| 36:05 | may not wanna add a complex you may wanna control and just add |
|
| 36:09 | by itself, right? So that's maybe a com maybe you just only |
|
| 36:13 | work with a defined me in some if you're studying things like that, |
|
| 36:17 | maybe not. But there's, there's for that in certain cases. |
|
| 36:22 | Um, it, everybody got Ok. OK. Uh Any questions |
|
| 36:29 | , about, about, about, this, any question about that. |
|
| 36:37 | . So let's go to this OK. So uh what's I already |
|
| 36:44 | up all the hints for you? , go ahead. Yeah, let |
|
| 36:50 | go back this one. Yeah. . So you're looking for, can |
|
| 37:00 | define which is a middle medium? means you don't have these things? |
|
| 37:08 | a complex medium. So, so that, that, that's why it's |
|
| 37:15 | , right? So why it's a , right? Heteros like you eat |
|
| 37:19 | like this, eat this and this ? You don't eat sulfur, |
|
| 37:23 | So that's so A B or C your choices? And the only one |
|
| 37:27 | there that's defined is b there's no nutrients in it. Yeah. |
|
| 37:34 | OK. All right. So look this medium. Uh what would grow |
|
| 37:40 | this medium? OK. So it's not a quicker question per |
|
| 37:43 | Just kind of mull it over and the ingredients. OK. Um Anybody |
|
| 37:49 | thought at this point before I roll the hints? Any thoughts, |
|
| 37:56 | OK. Well, um a what is missing? OK. What |
|
| 38:06 | missing? Anything missing here? Think the six letters, right? The |
|
| 38:14 | letter, whatever you call it, anagram. But uh here's a hint |
|
| 38:24 | do anything, see what's missing right? So what, what on |
|
| 38:34 | triangle? So something can grow on ? OK. Uh definitely can grow |
|
| 38:42 | this. There are things that can on this medium. So what |
|
| 38:45 | what side of the triangle contain members could grow on this got three |
|
| 38:53 | All right. Uh I'm, I'm buy me a new thingy after class |
|
| 38:59 | . Um You have three sides. see what's missing. OK. But |
|
| 39:05 | it can grow on this, It can grow on that medium. |
|
| 39:11 | which one, this guy, these these A b not a favorite |
|
| 39:19 | All right. A B or C of which root can grow on that |
|
| 39:25 | ? Mhm My phd work, I with these types. Which one anybody |
|
| 39:35 | on, what's the one of the we studied about that have their |
|
| 39:40 | portable nitrogen source with them? A fixation, right? So you can |
|
| 39:47 | on a medium without nitrogen. If can fix nitrogen, you get it |
|
| 39:50 | the air, right? 80% in air, right? Lots of |
|
| 39:55 | And that's how you, that's how study these nitrogen fixers as you grow |
|
| 39:58 | on medium without nitrogen. So that's we call enrichment culture. OK. |
|
| 40:05 | you are, so it's gonna we're gonna talk about differential and selective |
|
| 40:10 | well selective. And so there's kind a subtle difference between there, |
|
| 40:15 | And selective medium, which we'll talk next that you're purpose of purposefully adding |
|
| 40:24 | to it, to inhibit OK? types from growing and by doing that |
|
| 40:32 | favoring growth of other types, That's selective media. The um and |
|
| 40:39 | medium is you're not doing that. , you're, you wanna look at |
|
| 40:43 | particular metabolic type of micro. And you go OK, I'm gonna put |
|
| 40:49 | the things that it likes to right? And that's what you |
|
| 40:54 | So you're enriching for those types. ? So again, you're not adding |
|
| 41:01 | to inhibit things. You're changing the conditions to feed the supply of food |
|
| 41:07 | and nutrients that favor a particular You wanna look at whether it's nitrogen |
|
| 41:13 | or something else. OK. Um we talked about fastidious and so enrichment |
|
| 41:21 | pretty cool with that. Yeah. right, culture liquid. So we |
|
| 41:28 | about this already. Um like the and uh differential. So selective |
|
| 41:35 | Again, there are specific chemicals, chemicals. You you learned the lab |
|
| 41:40 | week. Um very common uh types media are those that inhibit uh gram |
|
| 41:48 | , favorite gram positives, vice versa Hibi gram positive, hi gram |
|
| 41:53 | OK. Uh differential media. Um can see color differences, basically usually |
|
| 42:00 | differences or clearing areas that represent some of hydrolysis of a component in the |
|
| 42:08 | differential medium. Like when it says can differentiate one group from another, |
|
| 42:13 | lactose fermenter, from a non lactose or those that can produce uh that |
|
| 42:20 | utilize uh sulfur compounds and produce a end product. Like you see over |
|
| 42:26 | on the right. Those that can't . Um Very often my knees are |
|
| 42:32 | for um uh wastewater treatment in the uh for those that make um drinking |
|
| 42:40 | . And so in drinking water, don't wanna have fecal contamination, |
|
| 42:44 | That's uh that's not water you wanna , obviously. And so there are |
|
| 42:49 | of that in the water, your coli other enteric that are indicators of |
|
| 42:55 | . And so you can use these of media to and so by |
|
| 42:59 | don't you know this E coliform uh one that can ferment lactose as in |
|
| 43:05 | negative rot, right? So uh immediate a lot of these media are |
|
| 43:10 | for looking for those types, So you can imagine the wastewater or |
|
| 43:15 | as you're making, drinking water with lot of stuff in there. So |
|
| 43:18 | wanna weed a weed out a bunch these things and look for those indicators |
|
| 43:23 | fecal contamination like E coli etcetera. . Anyway, so select the differential |
|
| 43:30 | this these kind of media you these aren't used, these are used |
|
| 43:35 | really in in um um diagnostic, know, clinical medical microbiology type diagnosis |
|
| 43:43 | . OK? You don't use these in like growing high cellules or stuff |
|
| 43:47 | that. These these kind of media used for differential media that is or |
|
| 43:51 | for more of these diagnostic type Um All right, we're gonna talk |
|
| 43:58 | little about quantitation now. OK. just to you probably already know this |
|
| 44:05 | , but just to give you a bit of perspective on how I keep |
|
| 44:09 | they grow fast. Well, how is fast. What are we talking |
|
| 44:13 | here in terms of numbers? So uh this is really just the |
|
| 44:18 | of doubling time. OK. Which guess you could also use in uh |
|
| 44:23 | , right? The uh the building of interest and blah, blah, |
|
| 44:27 | , so kind of the same very growth pattern. Um So here we're |
|
| 44:31 | with 10 cells, right? In scenarios. Uh So doubling time, |
|
| 44:37 | , We'll talk about that in a . So uh so you have 11 |
|
| 44:42 | has a doubling time of four One has a 15 minute doubling |
|
| 44:45 | So like an E coli has So what's the population size after 20 |
|
| 44:51 | ? OK. So everything upfront is same, right? 10 cells on |
|
| 44:57 | sides at 20 hours. What's so you, so this equation here |
|
| 45:03 | a basic one, right? So is always population size, population size |
|
| 45:09 | times zero versus T at some time the future. N is generation time |
|
| 45:15 | the nth, right? So um 20 hours, right? What do |
|
| 45:21 | have? How many generations in 20 ? OK. Well, the for |
|
| 45:25 | doubling time, one hour, uh generation every four hours. OK. |
|
| 45:32 | 20 hours, we have five OK. So plug in the five |
|
| 45:37 | ? 10 to 320 cells. Whoopee do the 15 minute doubling |
|
| 45:44 | which you might think, OK, less but how much of an impact |
|
| 45:48 | . One generation every quarter hour, ? Times 2080 generations, 80 versus |
|
| 45:54 | , you can already see that there's be a significant number. OK? |
|
| 45:59 | that, that doubling time, which is one way to look at it |
|
| 46:06 | um one cell to make two, ? More practical purpose purposes. It's |
|
| 46:12 | long for a population to double. . And 15 minutes versus four hours |
|
| 46:17 | a big impact. OK? And when we look at these, so |
|
| 46:23 | over on the right, right. here's a generation, right? Nothing |
|
| 46:27 | nothing complicated. Um So a cell , that's one generation. Of |
|
| 46:35 | bacteria can produce many of these very in, in what we call exponential |
|
| 46:40 | unlimited growth. OK? Uh I of it as they call it AJ |
|
| 46:46 | curve, right? Increases very Um And the generation time, |
|
| 46:54 | Is again, so whether it's one divided into two or more practical |
|
| 47:01 | population doubling, how long for that happen? OK. Now, uh |
|
| 47:05 | unlimited growth as we all know is something that's being forever a thing, |
|
| 47:10 | ? There's a limit to it because all based on any microbe in |
|
| 47:15 | experiencing a burst and growth like this due to an influx of nutrients. |
|
| 47:20 | ? Uh When that's not happening, is kind of, you know, |
|
| 47:25 | , nutrients are limiting. Uh but get an influx that can lead to |
|
| 47:29 | burst of growth. OK? That for a while. Because those nutrients |
|
| 47:34 | used up unless they're continually being pumped , right? Then they're gonna gonna |
|
| 47:40 | a limit and then they'll drop back again. OK? Just the nature |
|
| 47:45 | , of, of exponential growth, doesn't last forever. OK. |
|
| 47:50 | um and then again, right, number of generations, right? So |
|
| 47:53 | generations mentioned this before. Bacteria can this in 6 to 8 hours. |
|
| 47:59 | takes us 400 years to do 20 , right? So um so because |
|
| 48:05 | looking at rapid increases over, you , relatively short time period, you |
|
| 48:12 | of want to compress that scale. we use log to the base 10 |
|
| 48:16 | that. OK. Same as in right? You get uh ph |
|
| 48:21 | ph uh six to ph nine is ph units, but it's it's 10 |
|
| 48:27 | 10 times 10 like 1000 right? it's so we kind of compress, |
|
| 48:31 | have big number of ranges, you of compress using long the base |
|
| 48:35 | Same thing, it is kind of , right? So when we look |
|
| 48:41 | uh so we use that log the 10 in gross studies, we convert |
|
| 48:46 | to that to give us an idea how fast this thing is growing. |
|
| 48:51 | . The rates the inflection, the . All right. Is it is |
|
| 48:56 | very hi uh very high is it ? What have you? Uh So |
|
| 49:00 | use that, you know, use growth, can use the same bug |
|
| 49:04 | different growth conditions can give you different patterns of growth. Uh If you're |
|
| 49:10 | something, if you don't want to at the effect of an antibiotic or |
|
| 49:14 | , you have a new disinfectant, wanna check out, you may look |
|
| 49:17 | it this way with and without different , how it works. Growth, |
|
| 49:21 | cetera, all different ways to use . Um OK. So here's a |
|
| 49:28 | basic example. OK? Just, use it because you know this is |
|
| 49:32 | equation you can use. OK. And so if we have N zero |
|
| 49:38 | one cell, you wanna go. . Well, how many generations do |
|
| 49:42 | got if we go to four right? So remember um one |
|
| 49:49 | right? This divides now we got , the second generation, right? |
|
| 49:56 | we, of course, we have cells, right? And we can |
|
| 49:59 | that out easily by going um nt uh one, we started with |
|
| 50:07 | two to the nth which is right? Not surprisingly, it equals |
|
| 50:13 | . Would you do? Right? fine if you're dealing with like number |
|
| 50:17 | aren't so small as we get right? Which is what we do |
|
| 50:21 | a practical basis. This equation really have as much use for us. |
|
| 50:26 | need to get something a little bit that we can work with and figure |
|
| 50:29 | different parameters. And so that's why take this equation and we kind of |
|
| 50:35 | it and we use it doing a to base 10. OK. So |
|
| 50:40 | not gonna have to derive the In fact, you'll have the equation |
|
| 50:45 | on, on when the problem is , right? So you don't have |
|
| 50:49 | memorize that either. OK? It would help to know what the things |
|
| 50:53 | for like big N middle N. ? But the equation will be |
|
| 50:58 | right? So this is what we at previously. OK? So what |
|
| 51:02 | gonna do is to get this little generation time out of there and be |
|
| 51:07 | to use that to solve for different . OK. So very simply we |
|
| 51:14 | can uh go through with log to 10 right? On both sides of |
|
| 51:18 | equation, right? That's basically what's on here. OK. So if |
|
| 51:22 | remember how logs work, right? the the map of logs, so |
|
| 51:27 | log to the base 10 of two the nth um is the same as |
|
| 51:32 | this right, the end comes out here N times log the base 10 |
|
| 51:37 | two, you so for that is . OK. So remember this exponential |
|
| 51:43 | is like this power of two kind thing, right? Two to the |
|
| 51:48 | of power, right? And so kind of represent that in this number |
|
| 51:52 | value here. OK. So we it out to the end here |
|
| 51:57 | right? So this is our equation number generations equals log to the base |
|
| 52:03 | over the population. Size at T over what you started with. |
|
| 52:08 | . And so think of this 0.301 that two to the ends, |
|
| 52:13 | That exponential growth factor. OK. once we have this, you can |
|
| 52:18 | a lot of things with this. As we'll see, we'll do a |
|
| 52:21 | of problems here. OK. we very often are interested in the |
|
| 52:27 | , right? Putting how fast this occurring on a time basis, |
|
| 52:33 | Uh because that's what we're doing when growing cells, it's on a time |
|
| 52:36 | . And so we can add t that. OK. And so we |
|
| 52:40 | this k what's called the growth rate . OK. And so if you |
|
| 52:46 | E coli um well, a specific of E coli under the exact same |
|
| 52:53 | every time you should get the same rate, OK? When you begin |
|
| 52:59 | manipulate, you know, whether you nutrients or different nutrients or what have |
|
| 53:03 | or disinfectants, antiseptics, whatever you're to do, of course, that's |
|
| 53:07 | change right now under the optimum sticking with E coli under optimum conditions |
|
| 53:18 | they have all the nutrients they need there's nutrients they can use and you |
|
| 53:21 | the right temperature. Ph, et , et cetera, that, that |
|
| 53:27 | be, they'll be able to grow a maximum rate and that's what and |
|
| 53:32 | won't change. So every species has of that value under optimal conditions. |
|
| 53:37 | is as fast as they can And so every, every living thing |
|
| 53:41 | planet Earth has that intrinsic growth OK? This is as fast as |
|
| 53:46 | can grow. Even humans have OK. So, um but uh |
|
| 53:53 | anyway, when, when you're growing um especially in industry, you're gonna |
|
| 53:56 | , have an idea of what's the rate that we're getting here. |
|
| 54:00 | Ultimately, though the end game is many sales you're getting? OK. |
|
| 54:05 | how fast you get there is important . OK? Uh Because once you |
|
| 54:09 | in industry time is money. So generation time. So we can |
|
| 54:15 | this equation, right? Oops, gonna get to that in a |
|
| 54:20 | Uh You can take the equation here just flip it. All right. |
|
| 54:23 | remember generation time is um typically in usually is how it's how it's |
|
| 54:31 | Um And it's time per generation because bacteria they grow pretty fast. So |
|
| 54:35 | , that's why we use minutes and other time units. So um so |
|
| 54:42 | per generation, generation time. So let's, we're gonna use this |
|
| 54:46 | a couple of problems here. OK. So a bacterium has a |
|
| 54:52 | time of 40 minutes. OK. start, we're starting at five |
|
| 54:58 | OK. And we're log phase. you're not sure what that is |
|
| 55:02 | we'll get, it's, it's growing . OK. Basically. So how |
|
| 55:06 | minutes does it take to produce about cells? OK. Uh That's not |
|
| 55:11 | real strain. By the way, you're wondering, Houstonia Karen is not |
|
| 55:17 | , at least. So, um have five choices there. So let |
|
| 55:22 | open this up. So these are , these are the kind of problems |
|
| 55:27 | see again, there's examples uh on and um uh and, and they're |
|
| 55:38 | worked out and uh these will be , on cameras as well if you're |
|
| 56:20 | 100% sure, you know it, best shot, we'll go through |
|
| 56:24 | blow by blow, pause there for second. OK. Let's count down |
|
| 56:49 | . OK. Let's see what we here. All right. So I |
|
| 56:55 | of do this step by step So kind of setting everything up. |
|
| 57:01 | same, same question. OK. this is what we're looking for, |
|
| 57:05 | ? So N zero is five, going to 10,000. OK. So |
|
| 57:12 | how many minutes does it take? ? So if we uh if generation |
|
| 57:16 | is this right minutes over generation? . If we can figure out uh |
|
| 57:22 | we, and we're given the generation , right? Pretty much generation. |
|
| 57:27 | we can calculate the number of generations get from here to here, we |
|
| 57:32 | be able to do the math and um calculate that time, |
|
| 57:39 | So let's see. So we should something like this, right? |
|
| 57:44 | we'll figure out this and then multiply by the 40 minutes per generation, |
|
| 57:49 | ? So that cancels out generations and get minutes, right? So let's |
|
| 57:53 | how it happens there. So, boom. So 10,000 cells, |
|
| 57:59 | Is our NT five is our N and it comes out to be 11 |
|
| 58:06 | . OK? And so the mass 440 minutes, which is um let's |
|
| 58:15 | back. What's the uh forgot for minutes is I forgot my choices. |
|
| 58:20 | 40 minutes is um oh my my brain is cramping up. Um |
|
| 58:31 | my goodness do. Ok. What it say? 440 minutes? 440 |
|
| 58:44 | , 60 minutes to an hour, ? So that's about 567, a |
|
| 58:51 | over seven hours. Sorry, you need more caffeine, right? |
|
| 58:56 | so a little over seven hours, ? Um Every we're gonna do another |
|
| 59:02 | but everybody kind of get the set and how that's done. Ok, |
|
| 59:09 | here. Ok. Um that's another uh here. So Catholic and generation |
|
| 59:21 | , if 900 cells growing 15 hours over 3 million. Ok. Um |
|
| 59:34 | you're looking for generation time, like over generation? Ok. So again |
|
| 59:39 | just need to calculate that N value . Ok. Little n that says |
|
| 59:54 | , it should say two. sorry. No, might help if |
|
| 60:15 | do that. Here we go now can answer. Ok. Ok, |
|
| 61:13 | count down here from five 43. . Yes. If you answered 76 |
|
| 61:31 | , you are correct. So let's real quick go through this. So |
|
| 61:35 | just setting it up for 15 hours 900 minutes behind there. OK. |
|
| 61:41 | So starting with 900 cells going to many cells and zero to NT plug |
|
| 61:48 | the numbers, right? Almost 12 . Um So our generation time is |
|
| 61:56 | over generations giving us 76 minutes. . So again, these are the |
|
| 62:01 | of things you see on the You have an example of this on |
|
| 62:06 | uh canvas quiz this week. And a, there's a document called bacterial |
|
| 62:13 | problems in canvas as well in the two modules. So if you need |
|
| 62:16 | help with that, uh take a at that, of course, they |
|
| 62:21 | have any questions and it's possible you have arrived with the same answer in |
|
| 62:25 | different way. That's fine too. yeah, now it's gonna be pretty |
|
| 62:34 | kind of this thing here, either a generation time or finding how |
|
| 62:38 | how much time to get to this of cells, that kind of |
|
| 62:41 | right? So if you look at , the practice problems, it will |
|
| 62:44 | you kind of the five different examples , of what to expect. So |
|
| 62:49 | , it's just using the standard kind formula here. OK. OK. |
|
| 62:57 | right. So let's look uh switch and go into stages of growth |
|
| 63:03 | OK. So here's a question. . So bacterial batch growth curves. |
|
| 63:12 | while you're looking at this, um does batch growth mean? Uh |
|
| 63:18 | here we go. Let me use my prop. So you're doing batch |
|
| 63:24 | . All right, this is essentially batch. OK. So you would |
|
| 63:28 | media plop it in here inoculate and take measurements of some type to quantitate |
|
| 63:36 | growth. Usually it's using a spectra and optical density measurement and then you |
|
| 63:42 | a pattern like this. OK. And so what you do once you've |
|
| 63:48 | it, you're not doing anything else than taking a sample out to measure |
|
| 63:55 | growth. OK? And, and just following it all the way to |
|
| 64:02 | end. That's, that's it, batch growth. It's a batch of |
|
| 64:06 | you're following if you want to think it that way. OK. Um |
|
| 64:10 | there we'll see, you know, has its limitations depending on what you're |
|
| 64:16 | to do and we can, we um do other things, but that's |
|
| 64:20 | what batch growth is. OK. OK. So let's count down |
|
| 64:29 | Mm OK. So, e as consensus, let's see, uh which |
|
| 64:38 | is false. OK. So changes cell sign occur during phases two and |
|
| 64:43 | . That is correct. OK. changes occur in two and four. |
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| 64:48 | correct acclimation. That's correct. Most susceptible to penicillin, correct. So |
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| 64:55 | penicillin and the growth thing, faster growth like it, you |
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| 65:00 | assuming it's a uh um gram but even if it was a gram |
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| 65:06 | , you know, better chance of growing fast and it's not right. |
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| 65:11 | uh so e is correct or none these things are false. OK. |
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| 65:16 | let's just look at each of these uh briefly here. Um All |
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| 65:22 | So here's my batch right? We and then take samples and measure |
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| 65:32 | OK. Um And you can do the standard. So when, |
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| 65:38 | when these things grow, the liquid cloudy, right? And you can |
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| 65:44 | a spectra photometer to measure the level what's called turbidity, turbidity is |
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| 65:50 | OK. So it gets more turbid more cells grow and you can monitor |
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| 65:55 | . OK. Um Where there's no growth, it flattens out. |
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| 65:59 | But you can also at the same , take samples the same sample and |
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| 66:05 | an actual um what's called a viable . And there's a way to detect |
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| 66:10 | number of living cells that are in . OK? And that's the way |
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| 66:13 | get, also get the actual cell on on how it's growing. |
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| 66:19 | In any case, OK. Whenever first inoculate it, think think that |
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| 66:25 | the, you're the bacterium in that and you're popped down into this |
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| 66:30 | OK. So you have to consider did that come from? Uh How |
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| 66:36 | was I in there in my previous and medium? And um now I'm |
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| 66:42 | this new medium, um fresh brand , new uh just made. Uh |
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| 66:50 | there's gonna be subtle differences, uh not so subtle differences in things like |
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| 66:55 | , from where it came from where in now. Um, maybe 02 |
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| 67:00 | , maybe, um, medium maybe I'm in a different medium than |
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| 67:06 | was in previously. Ok. So that impact. So maybe it has |
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| 67:11 | turn on different pathways to use the it now has, uh maybe it |
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| 67:16 | to adjust the kind of this new . Ph I'm in now. Uh |
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| 67:20 | slight temperature changes. Uh So all these things mean that those cells |
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| 67:26 | are now in there aren't gonna begin exponentially. There's gonna be a period |
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| 67:30 | adjustment, so to speak. That's what we call acclimatization or |
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| 67:36 | OK? Uh That period of this lag phase. OK? Can |
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| 67:42 | course change, it can be OK? It can be longer right |
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| 67:51 | kicks in. OK. So there's number of factors that determine that, |
|
| 67:57 | , like I just said, gross , what am I? What was |
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| 68:00 | in? What I'm in now? . Um It can go both |
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| 68:04 | it can be shortened or lengthened depending those differences. The um uh you |
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| 68:10 | , going from a minimum medium to rich medium, right? That will |
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| 68:14 | speed up shorten like face because now being given lots of preform things to |
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| 68:21 | . Don't have to sit so long you know, express jeans, I |
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| 68:24 | get going quicker. OK. Vice , right? I'm on rich medium |
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| 68:29 | boom, I'm in minimal medium. now, I got to synthesize all |
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| 68:34 | express all these genes now to make pathways turned on so I can grow |
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| 68:39 | this stuff, right? That's right? So if these things uh |
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| 68:44 | many cells are going in like this cell going in, in the oy |
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| 68:48 | is it a million? Right? also influences blackface. So um so |
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| 68:54 | are all things that are are dependent each other. So in it, |
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| 68:57 | the industrial standpoint, when you want get this going pretty quick, the |
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| 69:03 | basically what you call these. So is your batch medium here, |
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| 69:08 | You have what's called the seed seed medium precede your batch medium. |
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| 69:15 | ? Seed medium, the batch, ? And it's in the seed that |
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| 69:21 | serves as Iran ocular. So if , if you're trying to do this |
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| 69:26 | to get lots of cells very you want both of those things to |
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| 69:28 | the same, right? Um And let that seed medium grow too |
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| 69:35 | right? You don't want that seed to be something that's gotten to |
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| 69:40 | right? And now you're using that the inoculate, right? Because it's |
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| 69:45 | that most of the cells in there dead, right? Very bad |
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| 69:49 | not gonna have enough stuff to even with right viable cells, right? |
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| 69:54 | , so you'd like to get a that's probably somewhere in this range, |
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| 70:00 | ? And use that because you're not much, much faster you're gonna have |
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| 70:03 | very quick transition to the next OK. So, um anyway, |
|
| 70:12 | , OK. So next phase, phase. OK. So log phase |
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| 70:15 | course, is the fastest growth, . OK. So it's, it's |
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| 70:22 | , it's now kicked into gear and are off and running. OK. |
|
| 70:27 | you may often hear the term uh mid log phase, which is kind |
|
| 70:31 | , it's gonna be the most active here perhaps. And very often if |
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| 70:39 | wanna measure an enzyme activity that that are making a particular protein slash enzyme |
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| 70:45 | want and you wanna measure its It's very often you do it there |
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| 70:49 | it's gonna be the most active. You may harvest cells at that point |
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| 70:55 | , you know, if you want metabolically active cells, you may and |
|
| 70:59 | , simply means to take this take this liquid and we're gonna dump |
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| 71:05 | in a tube and I'm gonna centrifuge . OK? And now all the |
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| 71:09 | are gonna fall to the bottom liquid poured off and then you got this |
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| 71:14 | , this wet paste, we call of cells and you can put them |
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| 71:18 | the fridge, you can freeze come back and use them later. |
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| 71:21 | that's kind of the process here when growing cells up and you wanna get |
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| 71:25 | out of them, you centrifuge That's what we call harvesting cells. |
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| 71:30 | ? Um OK. So log most metabolically active size, which so |
|
| 71:36 | is some solid size changes that occur this process. And remember in, |
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| 71:42 | log phase, the cell is gonna in this kind of state, |
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| 71:46 | The, the um all right. dividing state, right? Where they |
|
| 71:53 | be like this, lots of them , in that, in that |
|
| 71:56 | right? So, so they tend kind of, as they grow, |
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| 72:01 | get a little bit bigger and then you get the, as they get |
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| 72:07 | little bit too big, then that's sign that kind of divide. So |
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| 72:09 | gonna, that's where you're gonna, why they're the biggest in log |
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| 72:12 | OK? But then once you get , call this like late log. |
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| 72:20 | we're kind of getting into the tipping where there's not gonna be enough food |
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| 72:24 | sustain everybody growing at that fast rate . OK. So that's when it |
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| 72:30 | to slow down. OK. So stationary phase, OK? The |
|
| 72:36 | um the cell size actually decreases. . So now you're not, you're |
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| 72:44 | down to, you're not down to nutrients, but you are getting |
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| 72:48 | OK? And so now kind of stress responses kick in by the cells |
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| 72:55 | now it's just a matter of I need to survive, right? |
|
| 72:58 | the cells don't know they're in a growth phase, right? They're just |
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| 73:02 | to grow. OK. So they of then shrink a little bit size |
|
| 73:07 | down because if you're smaller, there's to keep up with, OK. |
|
| 73:11 | things like protein synthesis slows down to operate for essential things, processes. |
|
| 73:19 | . And um and so the goal from the micro perspective is OK, |
|
| 73:23 | me do these things to kind of , all right, maybe nutrients will |
|
| 73:27 | flooding in, I can start growing . Right. So it's kind of |
|
| 73:30 | kind of the um idea of what's on here, right. Survival, |
|
| 73:35 | . And um, so of if no nutrients come and they won't |
|
| 73:40 | we're in batch growth, right? not doing that, then death |
|
| 73:45 | right? And so there you're you're at zero now, zero |
|
| 73:50 | And so now, but it's they don't fall off a cliff all |
|
| 73:55 | once, right? So it's it's never this OK? Not that |
|
| 74:01 | a rate, it's a rate of kind. OK? Because everybody kind |
|
| 74:06 | , it's about accumulation of damage and kind of happens to everybody individually a |
|
| 74:10 | bit different. And so it occurs a rate. Ok? And so |
|
| 74:17 | that's all right, you guys you guys gotta go to lab, |
|
| 74:20 | in lab. So let me cut off there folks. But um but |
|
| 74:24 | , if you wanna, if you any questions, I'm, I'm here |
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| 74:27 | a bit. So please come up . Thanks. See you next |
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| 74:38 | Ok? Tell me, |
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