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BIOL 2321_Microbiology for Science Majors - 2321_10_19_23_CH 7_8
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00:00 So. Ok. Too much. . Oh, yeah. Right.
00:53 . See, testing, testing, , testing. Oh, there we
00:58 . All right. Well, low turnout today must be a all
01:05 something. I don't know the nice probably. I blame it on nice
01:10 . Um, ok, so exam tomorrow, uh, obviously the stuff
01:21 started last time and continuing today is three. Not on the exam.
01:28 , um, and I, I to double check that. I'm pretty
01:33 it is. CBB testing centers. . Ok. So you may be
01:38 to going to garrison, but check , it's gonna be CBB classroom and
01:42 building right over by Melcher before you to Melcher. But over there,
01:49 . Um, and I, I sent out the email yet. I'll
01:51 that. Uh, when I get and back to my office after class
01:55 send out an email, uh, . So you're, uh, I've
02:01 them, uh, the cost of that you're allowed to have calculators
02:05 So you shouldn't be hassled about Ok. So, uh, it's
02:08 a couple of problems. So, , but you'll be allowed any kind
02:13 , I don't care what kind just a cell phone, of course.
02:17 . So um so wondering why I'm this microphone. Sorry, holding it
02:23 this. Not on my body. have a reveal to make.
02:27 So we got big doings going on tomorrow. Saturday. What's happening
02:36 Big big one? OK. So be there if you see me,
02:43 hurl insults or throw rocks because Yes, yes, yes. Class
02:59 of 84. Yes. Yeah. if somehow U of H wins,
03:11 gets five points out of their, exam too, right? That's how
03:19 than I am, right? All right. Now they pulled me
03:26 said, uh we're not having so I don't want to show my
03:32 . OK. All right. Um . So uh OK. Gotta get
03:40 to uh not as much fun right. OK. Uh All
03:47 Let me tone down this microphone a here. All right. So uh
03:53 . So we're in chapter 78. . So this whole unit is the
03:59 of not the beginning, but we're about aspects of, of bacterial genetics
04:06 . OK. And so we're not into all the details of how DNA
04:13 and how, how protein uh expression in terms of the excruciating details,
04:19 ? Which you've, I'm sure you've that before. Uh But I'm more
04:23 on what's specific to bacterial oreal types and there. OK. So,
04:30 as I mentioned last time, we of did that overview? OK.
04:37 You know, it's, it's gonna if you can kind of remember the
04:41 because when we get into regulation, , uh it's all regulations all based
04:46 controlling the different steps of as you to DNA to RN A to
04:51 OK. So, um, so we'll go into a little bit about
04:56 and seven and eight is not the of those chapters. It's basically
05:01 these topics. OK? Um The and, and chapter eight is,
05:09 mostly um uh just the, the part and only as it relates to
05:17 , the Pro Kario type of that's it. Um Actually the
05:23 if you, if you looked at already, the stuff about OPERON um
05:28 gene organization, that's really mostly in 10, I think. But I
05:35 it's best to kind of seven and are kind of the, the time
05:39 , to explain that. All So, so we'll, we'll go
05:42 it today, but then you, be a bit of a, we'll
05:45 it again. We get the chapter . OK? But anyway, so
05:49 kind of what's going on here. , but I did uh forget to
05:53 questions we had last time. There's I always use uh as the introduction
06:01 to that, to that gene expression thing. And uh when I used
06:05 , when I used to teach intra , it was always a question on
06:08 exam when we, when we covered material. So, uh because it
06:11 much, um, we, we'll you right away whether you really understand
06:16 process or not. So let's look the question here. OK. So
06:21 is, let me pull it OK, I'll come back to
06:25 Let's look at the question. So here is our question. So
06:30 process of transcription translation is carried out a test tube. OK. Uh
06:35 a test tube is added. So have three sources. We have a
06:38 fish and zebra. OK. So hippo is giving us the Mrnatrnas and
06:45 fish is providing DNA zebra is providing RN A ply and any other necessary
06:53 , ribonucleotide and amino acids. So, um the question is assuming
07:02 new, new protein is made. . Uh The proteins of which animal
07:08 will be expressed. OK. um so read that carefully.
07:16 And uh this has been done, mean, this is, this is
07:18 back in the sixties. They do with a hippo fish and zebra.
07:22 they could um you know, it just goes to the universality of
07:26 process. You know, we all it the same way. OK.
07:30 , sorry, I'm blabbering on. . Uh All right, let's do
07:35 again. Here we go. Sorry that. Got carried away. All
07:41 . So, um yeah, you do this in a way call cell
07:46 extract. They call it. And so, and they did
07:51 um, to figure out where they the genetic code, they, they
07:56 like, um, different sequences of to figure out what code on went
08:02 , with Jimmy Acid. So, , so it's, it's something that's
08:06 fantasy. Ok. Mm. Is here born? Nobody was, nobody
08:33 alive in, here in 1984 were ? Ok. I think so.
08:39 be shocked. Ok. Um. . Mhm OK. Let's count down
08:54 . OK. Oh All over the . OK. All right. So
09:02 everybody agree that one of the animals has to be a fish. It's
09:08 be one of them, right? RN A protein, right? Uh
09:12 the other one? Why? Right. Yeah. Right. It's
09:20 hippo and a fish. OK. it's both of these. So
09:25 it's um so let's go back OK. This little review thing.
09:32 right. So if we look so we talked about phenotype and
09:38 right? And how that relates in of the flow of information,
09:42 So, uh traits being expressed, ? So genes are converted into uh
09:49 protein through the the workings of different A molecules, right? So we
09:57 for transcription, the RN A plier produce a RN A copy of the
10:02 . Think of it as the working , if you will uh of the
10:06 information uh that's then is translated into protein through the use of ribosomes transfer
10:14 A molecules Trnas and recognize a coon they bring an amino acid with
10:20 OK. And so, uh as see here, and so the ribosome
10:26 site is where the ribosome will bind a transcript. Uh We have like
10:30 , I'll talk about the, the UG and all that stuff here in
10:33 second. But we have um indicators the transcript of where it begins,
10:39 it ends and uh start code on one of the starter obviously. Uh
10:44 then the three base are the co that come after that. A UG
10:49 A UG itself is a code on . So, um and then uh
10:54 the way, so ribosome pro provide kind of the platform where all this
10:58 happening, uh not shown will be that recognize these coons, bringing the
11:05 amino acid and then we are producing polypeptide chain as we move down.
11:11 . So um the uh uh so look at bypass this to here
11:20 So, remembering that, that genetic table, OK, how we decipher
11:26 code on sequences? These are R a right, specifically from a
11:32 Um And uh it's redundant. And you, I'm sure you know of
11:38 there's 20 me lasses, but there's than 20 coons uh because there's repeatability
11:45 1234, coon, six coons for . So um the uh punctuation marks
11:53 you will start and stop uh beginning the transcript, uh what else.
12:00 um so looking at that plus minus , the um the um any sense
12:07 um relationship. So here is our the segment of DNA, this will
12:14 a part of a gene obviously. uh the sense DNA is the one
12:19 contains the actual information to make the . Um But as we know,
12:26 would be super easy if we could copy, copy the plus strand directly
12:31 a plus strand. OK. But , doesn't work that way,
12:35 The rules of complementary base pairing, . So the the we copy actually
12:39 antisense or negative strand that will then produced a plus strand. In this
12:45 RN A right transcript. So, these uh are identical, right?
12:52 the plus RN A and the plus strand are identical, obviously,
12:58 the, the T thymine are replaced U cells in RN A. Uh
13:04 uh anyway, so we've, we've that coding information to an RN A
13:09 . OK. And so you then the start code. So the orientation
13:14 this right at the beginning is the prime, it is the side that
13:19 , where it begins and then it to the three prime direction. So
13:23 here uh left to right. Um then you have a stop code on
13:27 some sort one of three that will the end and then each of these
13:32 for a particular uh amino acid. . So again, it's Trnas that
13:38 these coons through anti codons. Complementary anti codons and they, and
13:44 bring the appropriate amino acid with OK. So process, I'm sure
13:48 all, I'm assuming you've seen already to refresh your memory. OK.
13:56 But again, as I mentioned, the importance for us is that back
14:01 for a sec here, ear here um anywhere in this process, whether
14:10 at the level of the protein, uh transcript, the um the uh
14:19 of transcription. OK. So we block, we can block RN a
14:25 from even starting, we can um a, a made transcript and affect
14:32 we can have the Vibra zone and it in terms of its function.
14:38 you can affect the protein itself as as on top of that, you
14:43 manipulate the DNA and change that all things are done to control, control
14:50 process. OK? And multiple of can be working at one time.
14:56 . So it's all about. Uh I remember, you know, as
15:00 said, a bazillion times already this , the process you're seeing here on
15:06 page, right? That that's an energy requiring processes all over the
15:14 right? Whether you're copying DNA making protein, making a transcript, all
15:19 is building stuff, take lots of . So you don't want to waste
15:25 by doing this when you don't need . Ok. So that's why it's
15:31 tightly controlled, right? Your the bulk of your DNA in your
15:38 is likely devoted to some form of , right? Because you have a
15:43 more DNA involved, involved in that of stuff than you even do in
15:48 proteins in genes, right? Like 2% of your, of your genome
15:53 used to produce proteins. The other , right? Is gene regulation and
15:57 functions. OK? Uh Opposite of is most 90 something percent of bacterial
16:04 is protein coded genes. OK? again, whatever life form you are
16:11 genes is a big thing, Because you're not just gonna do those
16:16 when you don't need to. there's certain processes are pretty much on
16:20 the time. Like uh you certain metabolic process, you're, you're
16:26 eating and having to break down food get energy. So those kind of
16:30 involved in that may be on mostly the time. There's lots of other
16:34 that is OK? When you were zygote, right? You had a
16:39 of jeans being turned on to get from a zygote to a full grown
16:44 . OK? I don't think those are working anymore. OK? They're
16:48 , they're just not being expressed. . So anyway, um we'll get
16:53 into that in chapter 10. But um is, is after this
17:01 open another question, let me back uh any questions, right? So
17:07 terms of kind of this gene overview , I mean, I'm not gonna
17:10 you what does a ribosome do? . You're not gonna ask questions like
17:14 . So, um but you again, like I said, I
17:17 it's helpful to kind of re remember if you need to, you
17:20 I think make life easier. Uh But any questions, whatever.
17:26 . All right. OK. So look at this question. OK.
17:31 this is gonna lead us into this is so temperamental. Let's go
17:36 OK. So which of these terms all of the others? OK.
17:48 while you're looking at this, um will, I'm gonna show you the
17:58 system of how genes are organized, I'm not gonna be, you're not
18:02 be tested on it, but you to look at it for comparison.
18:16 . Yeah. So, all Counting down from nine. It's 5
18:30 . OK. OK. Uh of course. Genome is the big
18:37 . OK. Uh The order is gonna go genome one. What's number
18:46 ? No. So from biggest to ? Which one? Now going in
18:53 ? So biggest, the next the next biggest. What's below?
18:59 you know? Regulon? Yeah. act two and three. Anybody,
19:15 keep spitting it out. It's not operon. Yes. Three. Now
19:21 can say gene, OK. Then . OK. They the small,
19:29 . Um All right, we'll go obviously with opera and Regulon. I
19:34 think you've heard those terms yet, don't think. But, uh any
19:40 ? All right. So, genome and proteome. Ok. Uh Totality
19:48 DNA in any living thing is of , a genome and for most
19:52 it's going to be, you in us it's our 46 chromosomes uh
19:59 a, um, the bacterium, can be its chromosome but it can
20:03 associated plasmids. Ok. Um So it's what's it's the total amount of
20:12 uh the genes in an organism? . Genome. Hence, genome,
20:18 ? So tran transcriptome are basically the , the transcripts uh in the cell
20:25 any given time. OK. So because you take a, take a
20:32 , what do you got in terms transcript film and protein, right?
20:37 protein, of course, is all proteins present. So what would you
20:43 in terms of numbers? OK. larger transcriptome proteome? What would be
20:54 largest? Would you have more transcripts a bacterial cell or more proteins at
21:00 given time? Yeah. So if , if you measured how many Mrnas
21:09 in the cell at time X versus many proteins, what would be bigger
21:12 you think? OK. OK. , I'm not saying you're right or
21:21 . Um It turns out because I actually curious about that myself because I
21:26 my own idea that maybe it might transcripts but, and some missing information
21:33 is that RNAs in general, especially pro Caros last on the order of
21:40 . OK. Minutes. They, they're produced, they are translated and
21:44 kind of go away. OK. . Um So they're not super
21:49 So they don't hang around a So looking at, you know,
21:54 , is there any real data on ? So I looked, and for
21:57 one piece, I'll come back to . So one piece of data
22:01 this is a bacterial plankton, which guessing is cyanobacteria population. They tested
22:07 this and it's the same thing I've for like the numbers for E coli
22:11 others pretty similar. So that actually genes of the genome uh proteins are
22:17 most typically. OK. And so think I saw a number of like
22:23 for E coli, it was like no, it was 1800 transcripts.
22:31 ? That they've numbered and E coli about 4000 genes. So 4000
22:37 um uh 1800 transcripts and like over million proteins were quantitated. So,
22:45 know, it's just interesting. Uh was, I was curious about that
22:49 . So um anyway, but transcripts are transient, right? They're
22:54 and they go away um because you actually want it that way because
23:01 long as transcripts are intact, they be trans translated. OK? So
23:08 still may not want that, So a device kind of is
23:12 The transcripts go away after a If I need to make more,
23:16 just express more. Ok. you know, it's all, it's
23:20 controlled. Right. So, do need these transcripts? No, I
23:23 , I'm not gonna make more, I have already will go away.
23:26 right. So then I, I'll rid of making those proteins.
23:29 So, um, the proteins once they're made, um, have
23:36 longer lifetime, it, it varies type to type, they'll have a
23:40 lifetime in the transcript for sure. even the cell can, can,
23:45 degrade proteins uh that it wants to needed. Ok. But because you
23:52 , the importance of proteins in the , obviously to do the work of
23:56 cell, they, they're gonna be prevalent, right? And um uh
24:01 the genome of course, is the that's always there, right? So
24:04 not always expressing uh all the genes any given time, it's all about
24:09 . But anyway, it's not that gonna test you on this. I
24:13 thought it was interesting. And then , you know, uh but that's
24:18 of why the data, that's why looks the way it does,
24:21 So transcripts come and go kind of , uh protein is relatively more
24:25 OK. So um organization. So talked about this also back in chapter
24:32 , I think, kind of, know, the uh average car chromosome
24:39 if you will, maybe it's around million base pairs OK. That translates
24:44 around 5500, 3000 genes or that's kind of the average size.
24:49 always gonna be some on the upper lower range. E CO I is
24:52 , I think 4 million base Um The uh but then of
24:58 in terms of genome, it's not the chromosomes. So remember, proc
25:02 are haploid, right? One but you do have variants here and
25:07 some actually and and again, not majority, there's a few that have
25:12 in addition to that, they'll have linear some small linear chromosomes. So
25:15 are some kind of outliers, but most, it's like one circular chromosome
25:21 then maybe some plasma is associated with . OK. So plasmas are kind
25:26 uh extra chromosomal elements uh much smaller size. Um And, and kind
25:33 operate on their own. OK. uh the chromosome is tied to cell
25:40 , right? So the chromosome replicates to cell division, OK. Plasmas
25:45 of do their own thing. So not, they're not on that same
25:49 to that same uh uh process of replicating prior to sell. OK.
25:55 uh but we'll, we'll talk more plants here in a little bit.
26:00 . So OK. So read this . This is one of those before
26:04 afters, right? We're going to it again toward the end.
26:09 Um It, it might look at questions so far. Question question.
26:17 , good. OK. All So I'm gonna give me a few
26:21 to read through this again. You're see it again. So we're not
26:30 , once you answer it, well I can open this stupid
26:33 Ok. Sorry. Here we Ok. Ok. Do.
27:32 Mhm. Ok. Let's count down 10. Ok. 13.
27:47 Let's see what we got here. a picture. Ok. All
27:53 Um uh, I have, I a prep T A over in lab
28:01 I have to answer his text. . OK. Uh Yes.
28:11 Um Let's move on. We, gonna address all these things here as
28:16 go forward. All right. Um we're gonna look at organization of prokaryote
28:23 . OK? Um We're gonna look how the eukaryote looks only for comparative
28:30 . They're not gonna be tested on gene organization. OK? We're gonna
28:36 at it just as a comparison. right. So this term Cistron,
28:40 an old name uh for gene, don't really see it that much
28:45 but that's what that refers to. . So mono of course means one
28:51 means multiple. OK. So there bacterial genes for sure that are organized
28:59 one gene, one promoter. So , we haven't really talked about promoters
29:02 . OK. So the structure of gene, no matter what life form
29:09 is is um motor and then the gene being the, there'll be
29:17 , there'll be a um coding part the gene, right? The essential
29:23 preceding that is a promoter. The promoter is what guides the RN
29:28 plys to the front of the They're essential. You can't, you
29:34 have a gene without a promoter because the R A pli race won't even
29:39 because the promoter has elements in it , that the uh RN A pli
29:44 recognizes to put it right in front that coding information, right? Because
29:48 obviously what you wanted to do. want to code the entire uh protein
29:54 information. OK. So that's what the promoter are gonna be. Promoter
29:59 part of the structure. OK. , yeah, one promoter of one
30:04 , that's the mono Cytron, that's many uh pro genes can be that
30:09 , but most are in this operon . OK? And that's where you
30:13 one promoter and multiple genes associated with promoter. OK. So, um
30:21 so we'll look at that. So , that's that, that's that opera
30:24 . OK? Um Control is a part of it, obviously,
30:28 And so we're gonna look first here you carry on genes. OK.
30:35 , and some archaea have elements of as well. Uh But what's unique
30:41 the eukaryotes is their Exxon intron So the gene and, and you
30:47 out like you is has these express , exxons that are interspersed broken up
30:56 these intron sequences in between that you here. OK. So uh in
31:03 to that control elements, OK. Multiple control elements. So these involve
31:10 that bind and the binding promotes OK. So you have elements that
31:16 real close to the gene proximal and far away and by far away,
31:21 can be thousands of base pairs OK. And so that's what upstream
31:26 . Upstream always means in reference to gene you're talking about going toward
31:34 in this example, the uh right , OK. Uh downstream of course
31:40 passed beyond the gene. OK. so uh there's other sequences here,
31:46 was called a poly a sequence. This is transcribed into what we call
31:51 tail that is attached to the OK. Um The uh of
31:57 in the region where it terminates Uh So again, control elements and
32:03 of processing. So lots of RN processing occurs in eukaryotes. OK?
32:09 so the processing is necessary to produce transcript that, that has the introns
32:16 out. OK. So a translatable in eukaryote is only comprised of exxons
32:25 along with these elements of a called A tail. OK. And what's
32:32 a five prime cap. OK. so these elements, you have to
32:37 that transcriptions in the nucleus translations So the, the trans transcripts have
32:45 exit the nucleus. And so what them do that is having a cap
32:49 tail and that, that facilitates exit of the nucleus also, uh these
32:55 also affect um without them, they, they greatly enhance the stability
33:02 the transcript transcripts without a cap or , both rapidly are degraded.
33:09 The cap and tail need to be , not only for helping them get
33:13 of the nucleus, but also for them stable. And new periodic transcripts
33:20 comparison are much more stable than bacterial . E periodic transcripts can be,
33:26 , it varies, but the longest can last for days or months.
33:30 ? Until they're um till they go . So uh it depends. But
33:36 , so there's a lot of stuff on and you carry them basically,
33:39 ? In terms of transcription and OK. None of this do you
33:45 in bacterial, in a bacteria? . So um so in a
33:52 what we have is something like OK. So we've got, so
33:57 is gonna be the operon structure. again, some, you know,
34:00 , there are gonna be a number genes that have the uh one
34:04 one gene organization, but most are this operon structure, right? So
34:10 promoter and structural genes. So structural are the ones that uh basically are
34:16 the proteins that are being made. ? And so this one, in
34:21 example, we have three structural And um so what happens is uh
34:30 transcribe it as one continuous message. ? We call polycystic. OK.
34:38 this is all one continuous piece of A, OK. And so uh
34:45 , that then will be translated in this case, three different
34:49 OK. But this opera instruction structure that they're gonna code, they're,
34:57 gonna function as part of the same pathway. OK. So in this
35:03 , A B and C are enzymes catalyze this pathway here, W to
35:09 to Y to Z, OK. so uh very common in bacteria or
35:17 uh to organize their genes this right? Because now you can control
35:21 whole sect at one time. And that makes it a very efficient
35:28 . OK. We're gonna look at of these. One of them
35:32 is um how to sell will take we'll take lactose which you may have
35:43 lactose o on as well. two glucose and lactose. That's,
35:52 the lac opera. OK. So a cat pathway. It breaks down
35:57 , the products of that break down into glucose and lactose that can go
36:01 into like Colly and et cetera, ? That's one, the other one
36:06 one where we have a um uh called OK, Charis mic don't write
36:16 down. OK. Acid. And there's multiple pathways to get to um
36:24 the fan. OK. And so ruin the right trip to Japan.
36:31 . Uh OK. So to K , OK. So that's, and
36:38 mean it lasted, that's a anabolic . We're making it so that to
36:43 of set that in your brain. , right? Lactose opera is about
36:46 down lactose to get energy anabolic, tryptophan opera, which we'll look at
36:52 the opposite. It's anabolic, it's making crypto fan. OK? So
36:57 point here is that now you can that whole pathway in one shot.
37:04 . So again, makes for a efficient um system. OK?
37:09 um the OPERON itself, as you there is promoter operator, structural
37:18 That's, that's the OPERON. So what is the, the operator
37:22 for is a regulatory element? It in regulation of the process. And
37:28 how that happens is there will be regulatory protein uh somewhere upstream could be
37:36 away and the regulatory protein itself interacts the OPERON. OK. Operator,
37:43 . And uh the binding basically is physical block to transcription. OK.
37:50 it's what we call a transcriptional control . OK. We're basically, we're
37:55 allowing transcription to occur. OK? so the elements of how the trip
38:02 operon and lactose operon work is really what conditions bring that about.
38:11 And in lacto sauron, they're basically opposite in terms of what, what
38:17 that repressor. OK. And stops . So again, we'll look at
38:20 later. But um that's a very mechanism in uh proios. OK.
38:28 Now, uh among others, so again, all levels of expression can
38:33 controlled, but this is a very one. OK. And so the
38:39 for any questions about that for OK. So um OK. So
38:46 Regulon, so you've got the, the Regulon level is a, it's
38:52 step above opera. OK. So basically controlling multiple OPERON at one
39:00 OK. And an example of this we'll see an example of this in
39:05 nine in transformation and in uh there's uh in biofilm formation, this
39:14 OK. Um Where an example here about the control of I'll get back
39:22 that control of nitrogen in the right? So think of the different
39:28 uh a nitrogen source that a cell in can be used for what
39:34 Right. Well, you have an in amino acid. So for immi
39:38 synthesis, you have nitrogen in uh . So it's funneled into that.
39:45 you have um maybe just uh in for uh there's other connect containing molecules
39:54 the cell as well it's used So, you know, so it
39:57 different uses, right? So obviously important nutrient. So when it comes
40:03 different pathways may, may need it may not. So a way to
40:08 all that is through a Regulon. . So in that, then there's
40:14 be something common that all those operon are in that Regulon will respond
40:21 right? And that's what the Sigma is about, right? And so
40:25 talk about that toward the end, the sigma factor is what recognizes
40:31 OK? It works with RN A as you see here bottom. So
40:37 single factor works with RN A PLY bring it to the promote,
40:41 in front to the promoter, which in front of the coding part of
40:45 gene. OK? And so since this example, you see there are
40:53 well, they're showing you two, , yeah, two OPERON here,
40:58 ? And they're both controlled by that Sigma factor that affects both those
41:04 OK. So it can be more two opera, right? So however
41:09 are involved in the Regulon will tho those promoters will each respond to
41:14 same signal factor. OK. That's you can control the, these,
41:20 different operas involved in this kind of um process. OK. This example
41:29 nitrogen regulation. OK. So um Regulon um enable that to happen to
41:36 control of these different OPERON. Um Is there any questions about that
41:44 ? Yeah, proteins factors are, uh yes, yes. Um
41:56 So the S factor will, is , is a, can be a
42:00 controlling factor. Yes, but basically it is, it's for transcription,
42:06 the cell can also use it. Typically pro carrots have maybe 8 to
42:14 different Sigma factors. OK? And one, a general one that kind
42:19 is used for most of the genes the, in the cell, like
42:23 um uh DNA application and other things that. But then they have a
42:28 of others that are involved in specific of functions like this like to control
42:35 I think, I think moving a also involves a Regulon different operon.
42:40 different processes, the kind of do that way. So, so
42:45 one thing to think you may And E coli has 3000 genes,
42:49 that mean it has 3000 sigma No, it has, like I
42:53 , maybe 10 to 12 that work on these common promoters. OK?
43:00 yeah, go ahead. Um So factor uh something that isn't necessarily an
43:11 . Yeah, I would say um would say, I don't know if
43:17 protein is the right word. I say that by the fact that it's
43:23 to rec can recognize common promoters. ? Number one, the second factor
43:29 necessary to get transcription. OK? you might say the control factor comes
43:36 based, I would say more a factor may be a better word.
43:41 ? Because um the, the um if you like the, the example
43:48 the operator and something binding to it blocking transcription, the single factor is
43:53 gonna be able to do anything about . OK? That, that gets
43:56 in other ways. But the, the fact that they can coordinate with
44:01 promoters. So maybe think that as coordinating factor may be a better way
44:06 look at it. I think that sense. OK. OK. All
44:11 . Um But you have a OK. So uh let's look at
44:17 question. So we'll talk about plasmas . OK. OK. Um Oh
45:07 . Let's count out 10. Let's see what we got.
45:27 it's, it's c for sure. . Um So we'll look at plains
45:34 . So plains um as I mentioned , kind of their own, we
45:40 autonomous means kind of, they kind do their, their own thing.
45:45 . So uh since we discovered these uh 50 years ago or so,
45:54 we've since taken them into lab and constructed them for our own purposes,
46:00 our own plasmas. Uh I'm sure know, plasmas are one of those
46:05 that are in the gene gene, cloning, prominent DNA, right plas
46:11 essential to that. And so elements a plasmid. Um So what you
46:16 here is, is, is basically artificial plasmid but based on, based
46:21 a, an actual plasmid. Uh these things here refer to restriction
46:28 These are kind of the scissors that DNA, right at the, at
46:32 sites, you can open it and can insert DNA S uh fragments into
46:39 . Uh The gene cloning thing, ? Um Here you see this refers
46:44 antibiotic resistance, ampicillin tetracycline. Um So the plasma contains the genes
46:53 the cell has this plasma, it's to ampicillin or Tracy. Uh There's
46:59 origin of replication. That's what makes plasma autonomous, it has its own
47:04 replication. So it's not tied to division. Uh in order to replicate
47:10 can do it on its own and can have multiple origins of replication for
47:15 functions. OK. Um So copy , we'll talk about that. That's
47:22 in the context of how a cell hold on to a plain.
47:28 So it can be in high, be like uh 50 copies per
47:33 Low is basically 11 or two. . Um We especially see this in
47:40 nine, but the plasma can integrate the chromosome. OK. Uh Transferable
47:47 is where they're transferred. OK. different classes of plasmids. So the
47:54 to go from one cell to another is really through these types.
48:00 So we call an F factor class has the parts to enable conjugation.
48:08 . Now, these other features are , this resistance to antibiotics, heic
48:15 . So we a common one, might be back in 13, we
48:20 about um chapter 13 about aromatic This is the kind of things that
48:25 would see in a class is one those among others. OK. So
48:31 but you know an an F fact can combine so an F factor can
48:36 that or that, right? Having F factors is what makes it
48:41 it'll transfer. OK. So an factor can certainly contain an R
48:48 right? It can contain uh antibiotic . OK. So we'll talk more
48:53 that in chapter nine. But um in terms of replication, OK.
49:00 we're familiar with the usual um That's, that's how our, that's
49:07 the strand separation and, and then from each fork, right? And
49:16 your two daughter chromosomes. So um plasmas uh can do that,
49:25 But then they may have another ori them that allows for rolling circle
49:32 OK. So what that is is create a Nick, this is the
49:37 rep for replicate A I think what it does is it creates a
49:43 . And Nick basically means we are that kool aid bond in that um
49:50 structure. So remember uh you know three or five prime ends,
49:53 Three prime hydroxyl, five prime And so when we expose that,
49:58 you recall how DNA Climara works, looks for that three prime hydroxyl and
50:05 extends from it. OK? And from it using the, using the
50:12 this example, the inner strand is template, right? So it just
50:16 lines up right. Here's an I'm gonna put A T, here's
50:20 T I put an A and G blah blah blah, right? Complimentary
50:23 . And so as it continues along that inner strand, this example,
50:31 outer one is being displaced, being off, OK, as you see
50:37 . And so that can then be All right. So here's this.
50:43 here's now our this has been right? We're done here, this
50:49 one, the displaced strand and then be uh copied, right? So
50:56 primers, if you remember that you put a primer on there,
51:00 it extends the extends from that to it. So we end up with
51:06 two copies. OK. Now the circle application, we see that in
51:15 , it doesn't always have to only to happen in conjugation. But it
51:21 when, if a cell can do , that's when you see it as
51:23 . OK. And so where you'll it is so picture of these two
51:28 are cells. OK? So in , two cells come together and one
51:35 that, that plasma. OK? it's the rolling circle mechanism is where
51:41 see see that how, how it , right? So the the displayed
51:46 is shoved into the other cell. ? And so um uh and then
51:54 course, ends up copying that. we have a copy in each
51:58 So that's we'll get into the details that next time. But um that's
52:02 involved in conjugation. OK? Um questions about that. Yeah.
52:10 So inheritance. So number one plasmids not carry essential genes. So you
52:21 see a plasmid that has like the to uh like DNA polymerase on there
52:28 , to replicate DNA or, or for like ribosomes or, or you
52:34 , essential genes, right? Um things you see on there are things
52:41 antibiotic resistance, um uh other I mean, essential is kind of
52:51 , right? So it could be depending on if it's in the environment
52:57 having it ensures its survival, So, you know, because the
53:06 thing to remember again, goes back this thing about wasting energy and don't
53:11 things that waste energy is to have plasma and have it replicate, that's
53:17 energy from the cell as well that has. OK. So um especially
53:23 it's a high copy number class, that's a lot of energy to,
53:28 , to keep using, to maintain these plats. OK? Keep copying
53:33 , right? So you know, to hold on to a plans,
53:38 a decision, so to speak. ? That um you know, if
53:43 not being, if the genes on aren't being expressed, then it's likely
53:47 be lost. OK? At some . OK? Because it's just wasting
53:51 for the cell not benefiting it And so, you know, it's
53:57 eventually. OK. So um so how is it maintain? Well,
54:03 can, you can um insert can insert into the chromosome,
54:08 That's, that's one way to kind ensure it's it's uh retained.
54:14 And so think of antibiotic resistance, ? So here's kind of a really
54:19 example. OK. So, so selective pressure, right? Providing the
54:25 to force it to keep it, to speak. Ok. So how
54:29 you do that? Well, here E coli with a tetracycline resistance
54:34 OK. Here's one without, so means sensitive, sensitive to it.
54:40 the, if you grow it on with tetracycline, OK. Well,
54:45 course, it'll grow, it has resistance gene. OK. So that
54:50 is providing the selective pressure to keep , of course. Right. If
54:54 doesn't, it's not gonna live. ? But if it's uh sensitive,
55:00 course, it can't survive on their . OK? And, and the
55:08 the uh um uh without citrusy, you just grow in a medium without
55:14 , OK. The sensitive sensitive one course can grow. There's no nothing
55:19 it. Uh But the resistant one that grow on without citrusy course,
55:26 ? There's nothing inhibiting it. But thing is if you, if you
55:29 to maintain this resistant strain on a without tetracycline after several transfers,
55:40 It will go away, the plan just disappear. So you don't see
55:43 anymore. OK. So um so the thing, the selective pressure maintaining
55:50 obviously will ensure that it will um on. OK. But again,
55:56 a, it's a, it's, a we call it the business cost
56:01 benefit analysis, right? So is selective pressure there? That's the incentive
56:06 keep it. Is it not Well, maybe not? OK.
56:11 again, the, the loss of plant from a population isn't something that
56:16 happens in one generation, right? it, it can, it,
56:21 , it'll take several generations, but can certainly happen. Ok.
56:25 so the high copy, low OK. So one way to ensure
56:33 held on through successive generations is to have a lot of it in the
56:38 , right? So simply just by , you know, cell division,
56:42 it divides. So or like, , you know, pretty good chance
56:47 whatever plane it divides in it's gonna at least one on either side,
56:53 ? Uh So um that's a way ensure that it gets passed on.
56:58 . But again, that's, that's , that's kind of a price to
57:02 for having all these plats and using that energy to do it. So
57:07 , it, it's, it, depends, OK. So the um
57:16 low copy number. So for there can be looks kind of like
57:21 quasi mitotic spindle if you will. not that, but it kind of
57:27 resembles that. So, bacteria that these what are called par proteins that's
57:33 short for partitioning. OK. That um uh I I don't go into
57:39 whole mechanism here. There's multiple proteins , but the net result is that
57:46 these par proteins attached to the OK. And then begin to
57:52 And as they do, they basically each copy to opposite poles of the
57:59 . OK. So that ensures that it divides that each one now is
58:06 a copy of that plasma. So that's, that's something you see
58:10 cells with low copy number. And um it ensures, obviously that
58:17 occurs and each cell gets a copy that. OK. Um Of
58:23 that ensures um patches uh patches of plasma to the next generation.
58:30 Um Now, let's see. So any questions about that selective,
58:39 is this idea of selective pressure, ? Putting the the environmental condition there
58:45 um enhance the that the cells will that plan, whatever whatever kind of
58:52 it's carrying. OK. Um So our keto genomes, um there's
59:03 stuff that's different with bacterial types. the main, obviously, they are
59:09 , you know, don't I tend always when talking about prokaryotes, just
59:14 the word bacterial or bacteria, bacterial or that. Uh Of course,
59:20 including archaea in that I don't usually not explicit enough, but um you
59:25 , Procar so they, they, , of course, are prokaryotes,
59:28 ? They have a uh they lack nucleus, they have one chromosome.
59:32 . And so um and so what do have uh that's a little bit
59:38 is multiple AIS. OK? In chromosome, right? So, bacteria
59:45 have that they have one AI in chromosome. OK. And so a
59:51 can have multiple, that's just like , we have linear chromosomes. Of
59:54 , we have multiple origins. Um they do like rea, I
60:01 , sorry, like bacteria, they mostly protein coding genes. I mentioned
60:07 before. So the purple, you are no, is non coding
60:13 right? So you see in you carry out it's mostly purple.
60:18 . Um Green would be coding So not, not a lot.
60:23 ? Whereas up here on the top a prokaryote genome. Um lots of
60:29 , of course. Right? Because mostly protein coding. But remember um
60:34 know, there's a, a small of genes code for RNAs,
60:39 That's, that's the end product. always remember, kind of,
60:42 multi protein coding, but sometimes for genes, the end product is an
60:46 A molecule. OK. Um And that and uh DNA replication components.
60:55 things like uh DNA pli uh in , these have more similarities to periodic
61:03 DNA plier than bacterial uh in gene . Uh There are other elements there
61:10 arch are more similar to eukaryote than . OK? Like ribosome structure and
61:16 . But um anyway, so there's , there's certainly some similarities to eukaryotes
61:21 their genomes too. OK. Um right. All right. So the
61:29 part about eight I'm really going into on R ply race structure, specifically
61:35 Sigma factor uh structure. OK. uh a proc curia on a pli
61:44 have um parts that bind. Uh the sigma factor is the binding part
61:50 the molecule. It recognizes the promoter um of course, kind of like
61:56 puts it in front of the coding of the gene. OK. Um
62:03 beta and gamma, I'm sorry, and alpha sub units to summarize are
62:08 in kind of the synthesis of the . OK. And so uh there's
62:15 promoters have um specific elements to OK. Um We'll see an example
62:23 a couple different kinds but uh the 10 minus 35 is very common.
62:30 . In bacterial promoters and the uh factor. And so the S factor
62:36 not a, is not a permanent of the ply race. Signal factors
62:41 on and come off. OK. they guide, they're part of this
62:47 of the protein uh when they first to find, find the uh regions
62:54 bind to. And so they bind elements of the minus 35 minus
62:58 So that refers to 35 and 10 upstream from the start of the transcription
63:07 . OK. And so uh the factor is here. So it binds
63:13 then kind of starts looking for where's the, where is the uh
63:18 , these minus 35 minus 10. then that's where it will initiate transcription
63:24 the Sigma factor falls off. But it's free to bind another,
63:29 prelimerase. OK. So, um so that's basically how we initiate
63:36 OK. The uh OK. So terms of, of looking at multiple
63:42 these promoters, right. So a sequence is a sequence that's common to
63:48 of these promoters. OK. And , Sigma 70 is probably the most
63:54 Sigma factor in the uh cell. . So it recognizes these particular
64:01 And so these regions are, are to be a atat rich.
64:11 Specifically, because if you recall DNA , um GC base pairs have three
64:19 bonds. A TS have two. there's less energy to pull apart a
64:24 that's rich in A TS. And why promoters tend to be that way
64:28 that's where recognition and then shortly uh separation occurs. OK. So,
64:36 and so here you see in multiple minus yellow minus 35 minus 10,
64:44 the transcription start plus one. And so they have these common sequences
64:49 these two positions. OK. And you could, you know, uh
64:55 of the things you do, especially you're in biotech is to, if
64:59 want to uh affect expression of the is to fiddle with the promoter.
65:06 . Um So, a, a uh mutations, these are represent mutations
65:13 these pink positions here. The T C so changing the base there
65:18 to an A and A actually is down mutation means it means less
65:23 One that's up is one that increases . OK. And so, um
65:32 , uh and so that's a course interest, you know, if you
65:34 a, a protein, you discovered it's, it's gonna be the next
65:38 that's gonna make you a million right? Uh You need to make
65:42 of it, right? So you a combination of growing lots of
65:46 then you can also on top of uh increased expression of that gene as
65:52 . So both of those can are done and this is one way
65:55 do it is to find a promoter fill it with it. See if
65:59 get increased expression and and boom, can get where you need to get
66:05 . OK. Um Now, as said before, so promoter strength,
66:11 I'm gonna talk about that on the slide. So just hang on to
66:14 for a second. And so what really means means is the level of
66:18 , high or low, right? so uh not all genes you might
66:23 , well, all promoters must be , right? You wanna express things
66:26 at high levels. Well, you don't, right. So remember
66:30 metabolism, right? Uh in a , the the product of one pathway
66:38 be the reactive for the next one so on and so forth, you
66:41 , it can all be connected, ? And so the levels may may
66:45 all need to be high, some high, some are low to kind
66:49 coordinate the process, right? So so on purpose, some protein promoters
66:58 not so strong, they kind of with other promoters, right? For
67:04 expression. OK. Um Nevertheless, . So the, the, as
67:11 said before, you know, bacterium on average 3000 genes, but you
67:16 have 3000 sign factors, right? have a handful. OK?
67:21 the most common one being that Sigma . OK? Um But you do
67:26 others. So these are just examples you don't need to, you don't
67:28 to memorize this table. OK? just showing examples of other types of
67:34 factor. So here's one we talked nitrogen regulation, right, in the
67:39 of a Regulon earlier. So here's specific SA factor for, for those
67:46 in that Regulon, OK? They this one. OK? And here
67:50 for uh promotility chemotaxis that has a uh sigma factor. So again,
67:57 must also these must, that must be a type of Regulon as the
68:01 is involved in there, respond to particular sigma factor, right? So
68:06 an example. And so you also that although many of these are also
68:11 35 minus 10 that they recognize in promoter, the one for nitrogen control
68:16 recognizes a minus 24 minus 12. that's one that's a little bit
68:21 OK? So again, it kind set it sets it apart from these
68:26 types. OK? So again, know, that's what that's uh 12345
68:32 factors and that one's at six, ? And that's enough enough to kind
68:37 do what it needs to do. . Um uh OK. So what
68:44 strength and expression? OK. So we're going to see this when we
68:48 into regulation that there's um what we a basal level of expression.
68:54 So again, right, just basic , right? Orli sigma factor bind
68:59 a promoter and we have transcription, ? And so that can just what
69:05 see there, just those parts alone you what's called basal expression, which
69:11 basically means kind of low level OK? Um I mean, very
69:17 level, OK. In order to this ramped up, you typically involve
69:24 things. OK? So not just going to promoter, but things like
69:33 , transcriptional uh factors, transcription I'm sorry, uh these parts all
69:40 in. OK? And so there's example of those types of elements and
69:47 bind at the promoter and in doing , basically enhancing an end in,
69:54 um uh and especially in new periodic and maybe in some bacterial systems,
70:02 you'll, you'll involve like here's the , it too will kind of loop
70:08 and bind at the promoter as right, in response to these other
70:13 . And so this then creates this here, you circle it,
70:21 this right here is all happening at promoter and that, that now creates
70:25 site of high affinity for that It's like really it sticks to it
70:32 . OK? And the more it do that, the more expression you
70:36 , right, the binding is really affinity binding. Then you're really attracting
70:42 clime to that site and you of course, lots of oops,
70:47 of transcription, right? You have of transcription that way versus here where
70:52 kind of not so high affinity, get a little bit of binding and
70:57 , but this greatly enhances how much could be as much as a million
71:03 higher. OK? Expression, This could translate to maybe you
71:08 1 to 3 copies, right? can be a million copies,
71:14 And it can happen, oops three off of there, it can and
71:19 happens almost like that very quickly, ? Bacteria can ramp it up,
71:24 ? But again, it all depends , on the environment, what's out
71:29 , what's, what's, what's because things, these these factors are gonna
71:33 triggered to be produced, you by different types of conditions. And
71:40 but if it does, if it occur, then you can very quickly
71:43 up expression super quick. OK. We see that in the lactose
71:49 OK? You can go from almost like one or two copies to a
71:54 , you know, under the right . OK. And so uh
71:58 it's about enhancing that promoter. What we do to make that ply really
72:04 it, all right, and bind express, right? Any questions about
72:10 . OK. So I mean, in your own body, you
72:14 uh a lot of the things you um make cells do different things like
72:21 , growths, factors sold around your are basically have, have that function
72:28 as activators of expression, right? uh so that's what, that's how
72:33 cells respond uh to hormones and things that is getting into, to try
72:39 of different genes. OK. So any questions about that expression?
72:48 So um let's, this is just of a hold on. Let me
72:54 see something here real quick. Uh . You know what? This actually
73:01 a good fight to start. Next . It's kind of a summary.
73:04 uh wins five points on your exam . It's on record. It's on
73:10 tape and I win, I take points away. OK. Can I
73:24 bribery? Yeah. Yeah. Yeah. Here you see it from
73:41 am RN A but then like some the, I would say you can
73:47 get Mrnarn A. Uh Well, , the MRN A is the plus
73:55 . Oh, but it can come a minus. Yes. Yes.
73:59 minus what? It can only come a minus. OK. Oh,
74:05 it can only. OK. I mean, you can have a
74:08 RN A virus in facts. And who wants to make more copies
74:12 that, of its plus genome? gonna have to first make a minus
74:15 and then copy it. Right. . Yeah. Sorry. Uh,
74:28 , uh, yeah, you can tonight, uh, usually after seven
74:32 is when the kind of reshuffles things during the day and so seats typically
74:38 up. So check after seven Yeah. Uh, Monday or Wednesday
74:46 30 to 12 30. Yeah. , no, I'm, I'm out
74:53 the process. That's all on So, yeah. Oh, you
74:57 for? Oh, yeah. Yeah. Yeah. Yeah.
74:59 You can just stop by, stop , uh, you can do a
75:02 if you want but you can just by just first come first serve.
75:13 . Yeah. Yeah. Yeah. . Yeah, I know. 1
75:15 . 0, just come over to office. So I'll see you over
75:18 in a little bit. Yeah. a few minutes. Yeah.
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