Hi class, Dr. Jim here. In this chapter we're going to be looking at genetics and so we're going to spend some time looking at what DNA is, talk about transcription, talk about translation, and then the process of replication. So a lot of this will probably be a good review. If you haven't had a lot of this then hopefully you'll get something out of it. There are a few differences between bacteria and eukaryotic cells.
Bacteria again don't have a nucleus. The nice thing about bacteria is that they... don't, they only have one chromosome.
So that makes it a lot easier for transcription translation to take place. And so we don't worry about doing Punnett squares, we don't worry about dominant recessive in these situations. And we don't have to worry about a lot of different things that we worry about in eukaryotic cells.
So it makes bacterial genetics actually pretty easy, because all we have to concern ourselves is one gene, one protein. So in this section, I broke this up into two different sections. This is part one of chapter nine, and this is just going to deal with the basic genetics parts.
And then in the next unit, when we get into unit three, we're going to start with chapter nine, the second part, and we're actually going to look at regulation and how things are regulated. And so we'll talk about regulation, mutations, and a number of other things in that part. But for now, all I care about that you guys understand in chapter nine is, again, how things are replicated, and then through the process of protein synthesis, talking about transcription, translation, and all those different things. So let's get started on what we're looking at today. So the first thing we're going to look at is what is genetics?
And again, genetics is a study of heredity, and we're talking about that. We'll look at the structure of the bacterial chromosome. And if you remember, I talked about that it's a single piece, one single circular piece of DNA, roughly about four to five million base pairs. Sometimes it can be a little bit larger. Most of the time, that's about the average size.
And we'll talk about that and the difference between eukaryotic chromosomes. We'll talk about what are genes and what make up the genes. We'll also discuss genotype and phenotype.
Now, again, I told you we didn't have to know anything about pointed squares, but you still need to know the difference between what is a genotype and what is a phenotype. So genotype refers to the letters of the gene. Phenotype refers to the appearance of the gene.
And so we'll look at those things as well. We'll talk about the structure of DNA. We'll take a quick view again looking at what a nucleotide is, how DNA is arranged in the 5'-3'manner, and talk about why this is useful in the replication process.
So we'll talk about replication, semi-conservative replication, one old, one new strand, and how that gets done. And then finally, the last part of this lecture, we'll spend some time looking at transcription, translation, and the steps that are involved. Now, I don't go into a lot of detail about these things. I don't care if you know every single step of transcription translation. I just want you to know the process.
I want you to understand the process. And so what happens during transcription and then what happens in translation. And so I think those are more important keys than knowing every single enzyme and every single step. So even though I talk a lot about enzymes and things that are involved in these different things, I really don't care if you know all the different enzymes that are involved and what... what they do and how they do it.
I just want you to know the process. Understanding the process, you got to have to understand, you know, kind of see the trees from the forest type of thing. And I want you to understand the process and not get bogged down by the details of it.
Okay. So that's what we're going to look at today. All right. So again, I've talked about this. The genetics is the study of heredity.
And again, we look at genetics with that. We look at the transmission of biological traits. And so one of the things I always say in biology. First day of class is what is biology?
The ultimate goal of biology is to pass your genes from one generation to the next. And bacteria are no different. What they want to do is pass their genes on to their offspring.
And so this really is the heart of biology. This is what drives biology, getting that information from one generation to the next and passing those on to the next generation. And so that's really the essence of what genetics are. We'll look at the expression and variation of those traits and so some of the differences and we'll talk about expression again more in that next unit looking at the regulation and induction and repression and how they're turned on and turned off. In this case, we'll talk about the structure and function of the genetic material.
So we'll talk a little bit about how that's arranged and how the double helix and how those goes and then how it changes. Again, this is going to be in the second part. Uh, when we look at this in the next unit, talking about mutations and how this can affect things, especially in bacteria can create a whole new strain of bacteria based on just a few simple changes in what we look at.
All right. So again, just knowing some of the basics, the genome is the sum of the genetic material. And again, this can be chromosomes, the mitochondria, chloroplasts, and or plasmids.
So this would be for eukaryotes. This would be for prokaryotes. Remember, prokaryotes have plasmids. There's a small piece of the DNA. If you remember from back from chapter four, I was talking about these.
These are pieces of DNA that make the lives a little bit easier, but they're not required for the bacteria to survive. And again, when we talk about cells, we think of the genome as the DNA, the DNA structure and function. And then the genome of viruses can be DNA or RNA in this case. So that's what we look at. All right.
And then when we think of DNA, it's complex with proteins. And this makes up the genetic material to form chromosomes. And again, depending on what species you're talking about, you have the organism level.
And in this case, this is a fungus growing on a rotted log. You look at the cell itself. It has chromosomes on the inside, which tell it what to look like.
Inside the chromosomes, you have the DNA and that and where the genes are located. And then finally, when you get down to a molecular level, you're looking at the A's, G's, C's, and T's and how they're arranged. And that arrangement gives you variation of what something will look like. Okay, we look at bacterial chromosomes. Bacterial chromosomes are a single circular loop.
And so if we look at the prokaryote over here, you can see this is a single loop of DNA, double-stranded DNA. Roughly between four and five million base pairs again some are bigger some are smaller Depending on what we look at in its one loop we look at eukaryotic cells these tend to be linear pieces of DNA now It doesn't seem like they're linear because it seems that they're really compact in that and they are but these are linear pieces So they have a definite two ends to each Chromosome in each chromosome in these cases contain many many genes and so we're talking about the average eukaryote Those chromosomes tend to be about 100 million base pairs. So if you remember, I just talked about here, 4 to 5 million to 100 to 200 million here. So much larger and more of them in the eukaryotic cell.
Viruses have a very small genome. Viruses, if their DNA or RNA only have a few genes, they only need a couple genes in order to survive. And typically their DNA is anywhere from about... maybe 100 base pairs all the way up to a couple thousand base pairs.
And that's about the extent of what a virus is depending on how complex they are. Okay? All right. So we look at the chromosome. The chromosome is divided into genes.
And these are the fundamental units of heredity. And again, this is a site on the chromosome that provides the information for the cell. And then that segment of DNA contains a necessary code to make the RNA. which will then go make the protein. And so this is what we talk about when we look at protein synthesis.
And so the DNA is the blueprint, the RNA is the messenger, and the protein is the product. Okay. And again, we have three different types of genes. The first type in the majority of the genes are going to be for proteins.
We call these structural genes because you make a product out of these things. There's another set of genes that are the code for the RNA. And this RNA is either for the mRNA. or the ribosomes that are made or those subunits.
And then finally, the genes that control the gene expression, these are regulatory genes. So these regulate or turn things on, turn things off. And so these are going to be important. Again, when we talk about the operons and the induction repression, and again, that's going to be in the next unit when we talk about these different things being turned on and turned off. All right.
Again, even though we're talking about bacteria here, we need to know a little bit about genotypes and phenotypes and genotypes refer to again the letter or the organization of the gene okay so we concern ourselves with the genetic makeup the a's and t's now here is a eukaryotic organism this is a plant and again when we refer to phenotype and genotype the genotype refers to the letters and a lot of times refer to dominant and recessive now in bacteria we don't have dominant recessive you either have the gene or you don't type of thing and so it makes it a little bit easier We still need to know what genotype is. Genotype refers to the letters or the gene itself. The phenotype refers to the appearance. What does it look like? What does the gene do to produce what type of protein and what does the trait look like in the organism?
And so again, we talk about plants and that stuff. A lot of times we talk about petal color. In this case, the phenotype would be purple and this would be a dominant dominant versus a recessive.
Recessive would give you white. But again, in bacteria, we don't even have to worry about that because they only have one genotype, one letter for each phenotype, and that makes it very nice and easy. No opponent squares. All right.
So how are the sizes? So the smallest is a virus. And again, a virus is roughly between four and five genes.
Some are bigger, some can be more complex. And this comes out to about, like I said, about a hundred base pairs to about a couple thousand. The E.
coli, E. coli is the average size. Again, it has about 4,000 genes, and this equivalates to about 4 million to 5 million base pairs.
And again, this, if you pulled it out of the cell, would range to about 100 or 1 millimeter in length. So if you look at your millimeter scale, that's how big the genome is. Not very impressive in that stuff.
And I like this picture here. This shows you that the E. coli is actually releasing all its DNA out.
You can see how compact the DNA gets put into the bacterial cell. Now, if we compare that to our cells, our cells, we actually have 46 chromosomes compared to one, and we have about 30,000 genes compared to 4,000. So we have much more DNA packed into our cells. If you took all the DNA out of your cheek cell, that would range from one, from all the way from one end, all the way to the other. And most of you guys are under six feet tall.
So your DNA would actually be taller than you. And so that's the case. Now, if you're over six feet, then obviously that's not the case, but. It's pretty impressive by thinking if you just look at one cheek cell, and we'll look at cheek cells here, we did look at cheek cells, and you see one cheek cell. Inside that nucleus, you have six feet of DNA packed inside that cell.
Pretty impressive. All right, now DNA, again, the two strands are twisted in a double helix. I think everyone knows that. And the basic structure is the nucleotide.
And this includes the sugar, the phosphate, and the nitrogenous base. And again, In DNA, it's deoxyribose. And we'll talk about RNA here in a little bit, what RNA is.
But for now, just know deoxyribose. Deoxyribose is the sugar of DNA. And again, the bases are adenine, guanine, thymine, and cytosine.
And again, the bases are in the inside. Again, the sugar and phosphate makes up the backbone. The backbone is connected by covalent bonds.
So you see they're recognized by these solid lines. Inside, the bases are connected by hydrogen bonds. These are those weak interactions.
Those are the weakest bonds. And again, they attract to the other bonds, the other bases. Now, again, remember, with the bases, it's always A and T, C and G, or G and C, T and A.
Okay, so it always is that way. A's go with T's, G's go with C's. If you remember that, you always know what DNA coding looks like.
Okay? Now, again, this talks about this and how many... bases that you have. And so when we look at this, when you talk about G and C based together, they have three hydrogen bonds where A's and T's have two.
And so that's an important difference. And we can actually measure the content of someone's DNA based on how many G's and C's they have. And so the more G's and C's you have, the more stable the DNA is. And so typically most organisms have anywhere from 20 to 30 to maybe 40%, but there are some organisms out there that can have up to 60% G's and C's.
And so they are more thermodynamically stable technically than something that has only about 20% G's and C's. And that's because they have more bonds than that. And so that's just something to know. I'm not going to ask something like that on the test.
Now, something that I do want you to understand is anti-parallel. A lot of people have a hard time understanding this concept. Think of it as a street. So think of it as the street that you take to get into school. And so we go down that street and we drive on one side of the street.
The cars coming are coming in the opposite direction on the other side of the street. Well, DNA runs the same way. One side runs in one direction. The other side runs in the other direction.
And what does that mean? So let's kind of zoom in here on this DNA molecule and see what I mean by this. What they represent, these numbers here, five prime to three prime represent where the phosphate binds onto the sugar.
So you see the phosphate here. This binds to the five prime carbon here. which then binds to the sugar here.
You also have a three prime carbon right here, which will then go and bind to the next phosphate. If you look up here, you can kind of see how that works. Now on the opposite side, you can see everything is flipped and that the phosphate is actually up on the bottom.
The five prime is down here and the three prime is here. And so what you have is two opposite directions. So one is flipped. So it flips like this. So you kind of see it like this mechanism.
in this case. So one side of the DNA runs in one direction. So it points up, the other side points down, and that's what we mean. And so when we're talking about five prime to three prime, all that refers to is the carbon in which the phosphate binds to. And so again, just take a look at the slide.
So I give you the slides and the handouts. And so you can kind of look at it or look at it in your book. And this will kind of make a little more sense if you just kind of look at it for a little while and kind of understand it's all about where the phosphate binds. The phosphate binds to the five prime. And where is that phosphate and relative structure?
Okay, so one side's up, the other side's down. So think of it that way. Alright, so why do we care about the DNA structure? Well, really, even though it's a simple structure, it's very complex in the way that it functions. And so one of the reasons that this simple structure is so important is because it maintains the code.
And so when we reproduce, it's easy to open up the DNA, split into two strands, and build new strands on the inside. We call that semi-conservative replication. We're going to look at that in just a minute.
Okay, why is this important? It means that this is going to be guaranteed that the transcription, or I'm sorry, that the replication is going to be maintained, that you're not going to make errors. And then when you do make errors, you always have a fallback copy to go back on. You always have these old strands to go back on.
The other thing, the simplicity of the A's, T's, G's, and C's is that the arrangement of these things provides variety. And that's the structure of life. So we all look very different. We all handle things differently and do all these different things.
And this is based on our DNA. And so organisms are based on what their DNA tells them. And each organism has a different set of DNA.
And so again, even though we have similar A's, T's, G's, and C's, it's the arrangement of those A's, T's, G's, and C's that provides their variety. And so that's a really unique type of thing. So what I like to say is DNA is very simple, but it's ultimately very complex in the way that it does variety.
Okay. All right. So here's the concept check.
So in DNA... adenine is complementary for what, and cytosine is complementary for what. So I'll give you a minute and you tell me what those things are. Okay, if you said C, thymine, and guanine, you'd be correct.
So always remember, A's go with T's, C's go with G's. And if you remember that, you know one of the golden rules for DNA. You always know the bases that go together. A with T, G with C. All right, so the first thing we're going to look at is replication.
Now, I like to separate these out into two different components just because I think people get replication and transcription and translation confused pretty easily. And it is easy to flip around. The terminology is very similar.
They look kind of similar. You're dealing with DNA. And even sometimes I stumble up and say the term's wrong and you can always catch me on that. But what I try and do is keep these separate so that it kind of makes sense. So what we'll first do is look at replication.
Replication is making a copy of DNA so that you can split your cells into two. Okay. The next part is going to be looking at protein synthesis.
And so we'll deal with all the steps that are involved with protein synthesis. So for now, we're just looking at, let's split the DNA and build two new strands. All right. So again, what we like to think of is that replication occurs in both strands simultaneously, and this can create, creates a complimentary strand.
So what I want you to think about is the next time you unzip your coat, that's DNA unzipping. Okay. The zipper itself. is the enzyme helicase.
Helicase will unzip the DNA and open it up. And when that's open, now you have the room to build two new strands. And we call that semi-conservative.
So the idea is, is if you look at this diagram over here, is that you can see this is blue and this is purple. The blue represents the old or parental strand. The purple represents the new strand.
And this is how it's built. Same thing on this side. You have the old and then the new.
And so this is a semi-conservative process. What this means is that you always have one old strand and one new strand. And the reason for this is if you make a mistake, you can always go back and fix the mistake using the old strand. And so that's the reason why we always like the semi-conservative because it opens up and you build two new strands and you have a way to go back and fix it. Okay.
All right. So in DNA replication, we are making an exact duplicate of DNA. And again, this involves about 30 enzymes. And I don't want you to know.
all 30 enzymes. We're going to talk about a few of them here. And again, really the one that I really want you to know is helicase because helicase is really important in unwinding the DNA.
If you know DNA polymerase 3 and all these other things, that's fine. I'm not going to ask you that on the test, but those are some of the proteins that are involved in the enzymes that are involved in opening and making copies of the DNA. So remember, this is all about making a copy of DNA so you can split your cells.
So again, in bacteria, this begins at the origin of replication. So this is kind of the site where the zipper would stop. So think of it like in your book bag where you have one spot where the zipper will open and close and maybe that zipper goes all the way around like in a luggage and you have that zipper that can zip the whole top off.
That's kind of like the start. That's the origin of replication. So that zipper will go and open up the DNA at that site.
And again, the first thing that happens is helicase will unwind the DNA. So what happens is helicase, the protein binds on the DNA, breaks the hydrogen bonds, and then works its way around. And then you have another one going this way, and eventually they'll meet in the middle, and now you have the two new strands that open up. Okay. Again, at the start, you have this primer, the primers added to the DNA.
And so now you can build the new DNA strands. So the primer is represented in green here. And again, the DNA polymerase binds on, builds it in this strand, builds it in this strand. You can see back here, these are shorter little segments.
And again, that's called the lagging strand. We'll talk about that in just a second. This is because of the orientation, the five prime three DNA can only be built in a five prime, three prime. So the opposite side has to be built in an opposite direction. And so that that's one of the tough things about DNA.
And so I really don't like to get bogged down. If I was teaching a molecular class, then I want you to know these different things. But since this is a microbiology class, I really just want you to understand the process of these things that are going on. Okay. And again, I talked about the leading and the lagging strand.
The leading strand is the strand that gets built in one direction. the lagging strand are these short little segments that get built behind it and so you think about the leading strand this gets let out from the origin whereas these lagging strands are coming up from behind so that's kind of how i think about it and it's a short little segment of dna as we look at those things okay and again then you have other things that will take these parts out and then eventually it makes one continuous uh piece of dna and so this is how the dna polymerase one and these ligases will remove these primers and fill these in and build the new DNA so that the lagging strands get all into one piece of DNA as well. Okay. And this just kind of shows you again, the leading versus the lagging.
And so again, you see how these things, here's the helicase, it opens it up. And then you have the lagging strand where it makes these smaller little pieces and this should be leading strand. This is not correct over here. So this should be the. leading strand.
You can see one strand gets built in the correct orientation. The other one has to get built in little pieces. And I have a video to show this here. So if you're not getting it quite offhand right now, watch the video and I think it'll make a lot more sense. Okay.
And so again, what happens is that you open the DNA and think of this unzipping, you're unzipping the zipper. And so you form two circles and eventually an unnix and eventually you get the two new daughter cell DNA. Okay.
And so that's really the process. And again, I'll show you a video here and it'll make a lot more sense as we go along. OK, here's the video. DNA replication begins at a specific point in the DNA molecule called the origin of replication site.
Initially, the enzyme helicase unwinds and separates a portion of the DNA molecule. after which single-strand binding proteins react with and stabilize the separated single-stranded sections of the DNA molecule. The enzyme-complex DNA polymerase engages the separated portion of the molecule and initiates the process of replication.
DNA polymerase can only add new DNA nucleotides to a pre-existing chain of nucleotides. Therefore, replication begins as an enzyme called primase assembles an RNA primer at the origin of replication site. The RNA primer consists of a short sequence of RNA nucleotides, complementary to a small initial section of the DNA strand being prepared for replication.
DNA polymerase is then able to add DNA nucleotides to the RNA primer and thus begin the process of constructing a new complementary strand of DNA. Later, the RNA primer is enzymatically removed and replaced with an appropriate sequence of DNA nucleotides. Because the two complementary strands of the DNA molecule are oriented in opposite directions and the DNA polymerase can only accommodate replication in one direction, Two different mechanisms for copying the strands of DNA are employed.
One strand is replicated continuously toward the unwinding, separating portion of the original DNA molecule, while the other strand is replicated discontinuously in the opposite direction with the formation of a series of short DNA segments called Okazaki fragments. Each Okazaki fragment requires a separate RNA primer. As the Okazaki fragments are synthesized, the RNA primers are enzymatically replaced with the appropriate DNA nucleotides, and the individual Okazaki fragments are then bonded together into a continuous complementary strand. Okay, and so again, I think this is a good little video just to show you, and I didn't really talk about these, but again, here's the leading strand.
The leading strand is made continuously and the lagging strand has to be built into the little segments. And we call these segments the Okazaki fragments. And so that's the difference here. And again, I'm not going to ask what is an Okazaki fragment or anything like that, but I do want you to understand the process. And so again, this is the unzipping of the DNA.
Helicase is responsible for unzipping. And then you build the strands 5'to 3'. And again, you make two new strands from the two old strands. And we call that semi-conservative replication.
Okay. So now we're on to transcription. And so transcription is involved with making proteins.
And so this is done all the time. Replication is only done when you want to divide the cells. And so replication only happens once and then you divide the cells and then that process starts over when you want to divide the cells again and again and again.
Transcription takes place all the time. And so that's one of the important differences between these things and what you want to think about. Replication is only going to happen when you want to divide the cells. where transcription is going to take place over and over and over again, because you need proteins in order for your cells to survive. You need to make enzymes.
You need to make some proteins for structure. You might need them for support. You might need them for building.
You never know what you're going to need those proteins for. And so that's why transcription will take place all the time. So that's one of the big differences between these six.
Okay. And again, this is the application of the genetic code. So we're going to talk about how DNA is converted to RNA.
And when that DNA is converted to RNA or changed, we call that transcription. And so it's making a copy of the DNA. And so it's probably not the best that you use copy because we just talked about making another copy of DNA.
But in this case, we're making a DNA to RNA copy. So what I like to refer that to when you're thinking about it, think about if you missed class and you wanted to copy someone's notes, you're going to have a little bit different handwriting and you're the tRNA or you're the mRNA. making a copy, word for word copy of someone else's notes so that now you have a copy that you can take home with you.
And that's essentially what's happening. We call that transcription. The next step is where you change that RNA into protein language.
And so there are two different languages. If we talked about the two molecules, we talked about the two different biomolecules, you have nucleic acids and you have proteins. Nucleic acids are read in one fashion. then proteins are read in a different fashion and you have to go from two different types of languages. And so we call that process translation because you're translating the RNA language to a protein language.
And so this is taking that genetic code and converting it into protein code. Okay. And so that's kind of how I refer to it.
So think of it as, let's say you have a roommate that doesn't speak any English, and now you have to translate your notes that you made a copy from your classmate. And now you're making... in English to Spanish or English to French or whatever language, you're making a copy of that so that now that they can understand it as well. And so that's the process of protein synthesis. First you make a copy and then you translate it into the language so that the proteins can be made correctly.
Okay. And we'll look at all these different steps as we go along. We call this the central dogma of biology.
And so this is because this is so important. for organisms to live. If you don't make proteins, you're not metabolizing, you're not going to function. And so you need to go and take the blueprint of DNA, convert that to RNA, which I call the worker, and then that worker will build the product, which is the protein. And so again, this kind of just represents.
And so what we think of it is that you have a triplet of DNA or RNA that codes for specific amino acids. So here represents an A, G, and T. Okay. The protein's primary structure is the amino acid organization.
So this is where you go back to protein, proteins and primary, secondary, tertiary. I told you everything builds. And that primary is the type of paper you use.
Okay. Remember when we were building the paper airplane and this is what it looks like. And so that DNA tells you what the protein is going to look like by telling you what amino acid is going to go in what order. Okay.
And so you think of it that way. And again, the proteins determine the phenotype and then the DNA is the blueprint. It tells you what the cell is going to do, how it's going to function, what it's going to make, and all those things. So the DNA is the important. This is like the library to tell you what that cell is going to be, how to build that cell, okay, and what gets made by that cell.
And so that's the important thing right there. All right, so first we need to understand what is RNA. And again, probably most of you know the difference between DNA and RNA. But again, I just want to talk about the differences. And again, it's a different sugar.
So the sugar is different. Instead of deoxyribose, we have ribose, and that makes up the RNA. The other main difference is that instead of thymine, you now have uracil.
Uracil is the other base. You still have adenine, guanine, and cytosine, but instead of T's, you now have U's. And so one of the ways you can always tell if a sequence is RNA or DNA is just look for the U's. Do you see T's? that should think in your head, that's DNA.
If you see use that's RNA. So whenever you see use, think RNA. Okay. And again, we have some different types of RNA, the three that we're really concerned with and what we're going to be talking about in the class is messenger RNA or mRNA. And this is going to make the message from the DNA.
So it's going to transcribe the DNA code into RNA. And then that RNA is going to come out and then bind to the ribosomes so they can be made into the protein. The tRNA is involved with bringing an amino acid to the party.
So this is like the guy that's in charge of bringing the beer to the party. This guy is going to bring the amino acid so he can build the protein. The RNA is part of the ribosome. And so we really don't talk much about the ribosome except the ribosomal RNA, except for the subunits. And so I just want you to be aware of some of that subunit is RNA in there.
And so that's called RNA. And then the primer, again, this is for replication. So we won't even worry about the primer for today.
So just think of these three, the M, the T, and the R and what they do. Okay. And again, I've just talked about the different ones. So messenger RNA carries the DNA message.
And so what happens is you make a copy from the DNA and that is the transcription part. That transcribed part then is then chain or taken to the ribosome where then you make the DNA or make the protein structure. Okay. So think of. RNA as the message going from one place to the other.
The transfer RNA is responsible for bringing amino acids to the party. And again, it has an amino acid attachment set at the top, and then it has the anticodon. And we'll talk about codons here when we get into translation in a little while. Now, the reason why it's called tRNA, if you look at it, kind of looks like a T.
And so that's how we refer to it, even in the... molecular structure, it still looks like a T. So it kind of looks like an uppercase T.
This kind of looks like a lowercase T. And so that's where really the tRNA comes from because it kind of looks like a T in the structure. Now, the last one is the ribosomal RNA. This is the RNA.
And again, we're talking about the subunits. And remember, the difference in prokaryotes is that they have a smaller subunit. They have a 30S and they have a 50S, which makes the 70S. In eukaryotes, they have a 40, a 60, and that makes an 80S. And again, Don't add because it's all about things spinning down.
All right. So let's set up transcription. And this is the first stage of genes expression.
So instead of DNA polymerase, now we're talking about RNA polymerase. So you got to think we're going to make RNA. We're making an RNA copy. So what makes the RNA copy this RNA polymerase? What it does is it binds to the DNA upstream of where the gene is.
And we call that the promoter. The RNA polymerase starts to add nucleotides. builds it in a 5'to 3'direction. And this is going to make a little more sense as we go along.
Again, instead of having T's, when you have an A, you put in a U, the opposite. And again, instead of the T's, you put in the U. And then finally, when it ends, the whole thing falls off and now you're ready to start translation.
And mRNAs tend to be anywhere from 100 to 1200 base pairs, depending on the size of the gene. And again, some genes are longer than others, and that all depends on the size. And typically roughly around 500 base pairs is about your average size of an mRNA.
Okay. All right. So let's see some pictures and see if this makes any more sense.
Okay. So the RNA polymerase is involved with really kind of the unwinding and building of the strand. Now we have two strands of RNA here. We have what is called, or I'm sorry, of DNA.
We have what is called the template strand and the non-template strand. The template strand means this is the strand that's going to be used as the template. how you build your RNA. Okay. And so it is going to look basically the opposite of what you have on this strand, because these are complimentary in that stuff.
So what I want you to think about on this is when you build your mRNA is that it's going to be exactly the same as this lower piece of DNA. The only difference is instead of having T's put in, you're now going to have U's. And so this is building it in a five prime to three prime manner.
So on the test, I will give you a sequence. then that sequence will be already the DNA five prime to three prime. All you have to do to get the mRNA is just change the T's to use. Okay. Now you've probably done this in other classes where they really try and confuse and everything else, but let's keep it easy.
Let's say we have our five prime to three prime. We flip it. And all we do is say, take that five prime to three prime and just change out the T's and add use. Okay. So you don't have to do anything else.
So I'll give you the sequence five prime to three prime. The mRNA is five prime to three prime. All you have to do is change it from T's to U's and you'll have the answer correct.
Okay. And again, I had videos for all these things, so it'll make a lot more sense of how this goes along. Okay.
So here's the video here, watch the video and I'll, I'll come back in and show you here in just a second. The synthesis of messenger RNA is called transcription. Transcription begins when RNA polymerase recognizes and binds to the promoter region on the double-stranded DNA molecule.
A particular subunit of the messenger RNA, called the sigma factor, participates in recognition of the promoter region. Soon after transcription is initiated, the sigma factor dissociates from the RNA polymerase. RNA polymerase moves along the template strand of the DNA, synthesizing the complementary single-stranded messenger RNA molecule. Synthesis is in the 5'to 3'direction, with new nucleotides being added to the 3'end of the growing messenger RNA molecule.
As the RNA polymerase advances along the DNA, it melts a new stretch of DNA and allows the previous stretch to close. When RNA polymerase reaches a specific sequence of nucleotides on the DNA called the transcription terminator, a hairpin loop structure forms in the messenger RNA, causing the RNA polymerase and the messenger RNA to dissociate from the DNA. Okay, so I really like this video because it really shows you exactly what's happening here.
And again, remember, 5'to 3', this is one direction, this is in the opposite direction. And so RNA has to be built 5'. prime to three prime. So it's going to use the bottom strand, which is the opposite strand.
So this is what I was talking about when you have the top strand, this is the top strand. And so let's say the strand was ATGC. Okay.
Let's just keep it easy. Well, the mRNA is going to be the exact same strand, and this would be AUGC because again, there's no T's involved in that. So the only difference between this in this is that instead of having T's in this, now you're going to have U's, but the rest of it is the same as this string. So what it does is it reads the bottom and rebuilds the top, just building it basically an exact copy, except replacing the T's with U's. So hopefully that makes sense.
If you still need help understanding that, I would be happy to go over this again in lab with you guys so that you just kind of get an idea or even after class just to kind of get it. Because this is kind of one of those things that it's just... Once you get it, you'll understand it, but it's sometimes hard to just grasp the first time. So watch the video again if you need to, and kind of go through it and say, oh, I finally get it. So, but if you do have questions, let me know.
All right. So the next step after you make the mRNA is now you need to translate that mRNA from nucleic acid message to protein message. And so you want to go from nucleic acid to protein. And so that's the important step. And we call that translation.
So you're translating it from one language to another. And again, translation is the second stage. And again, all elements are needed to synthesize protein are brought together at the ribosome. So the site of this is at the ribosome.
Now, prokaryotes, this is going to be in the cytoplasm, just like where the DNA is. In eukaryotes, it's also in the cytoplasm. That's where ribosomes are. The difference between the prokaryotes and eukaryotes is that transcription will take place in the nucleus.
of the eukaryotes, whereas in prokaryotes, there is no nucleus. So these can actually take place simultaneously. So first you have transcription.
And as soon as transcription is made a little bit of mRNA, you can already start translation. And that's kind of one of the nice things. And again, there's five stages to this process.
We'll go through them. And again, I don't care if you memorize the different stages and that stuff. I just want to show you these different things of how this is done. And really for you just to understand that.
the process. Again, don't memorize the enzymes and things like this. Just kind of get an idea or grasp on how this process actually works. Okay. All right.
So the first thing you have to understand is that basically what that mRNA is, is a long sentence of three letter words. Okay. And so now what we're interested in is all these three letter words.
And if I gave you this assignment and said, tell me how many three letter words can you make using just a, you, G and C, you would sit there and go A, A, A, A, U, A, A, U, U, and so on and so forth. And you could go through and come up with a total of 64 words. Okay.
And so we have 64 words that mean a number of different things. Now in English, we have lots and lots, we have more than a hundred thousand words. And so we have to have these big dictionaries to understand what all these words mean. And sometimes we really don't know what we're saying. And so we think we know what we're saying.
We don't. but that's another story. Okay.
In this case, we have 64 words, so we can fit all these words and what they mean on one page. And that's kind of nice. So think of it this way, what these are is a collection of three letter words. Okay. We have these three letter words and all these three letters mean the same thing.
So you can see here, this is called a codon usage map. And so this is the, this is the dictionary to tell you what these three letter words mean. So I know what U U U means, it means phenylalanine.
UUC also means phenylalanine, where CUU means leucine, or AUU means isoleucine. And so I know now what the definition of that word is based on using this code on your system app. What this refers to is this three-letter word tells you what amino acid goes in that spot.
And so this is a code that is found in all organisms, starting with bacteria, all the way to humans. We have the same definitions in our DNA as well. So you could go in your DNA.
and figure out all the definitions based on those three letter words. And this code is redundant so that you have more than one amino acid for each letter. Okay.
Or for each word you have each word is there's more amino acids because there's only 20 amino acids. And so what that redundancy means is that you have number of words that mean the same thing. And so if things get changed by mutations, it really doesn't affect the DNA too much or affect the protein too much because you have this redundancy in what they mean.
And so hopefully that makes a little bit of sense there. All right. And so now we want to interpret the DNA code. So again, before, when we were talking about DNA to RNA, we didn't really care about the three letter words. All we cared about is we wanted to make sure that we got the, the DNA letters to RNA letters.
Okay. And so we made the the DNA and made the RNA. And so here is our RNA.
We know it's RNA because it has U's and no T's. And so we just cared about the letters. Now, when we go from RNA to protein, we're concerned with the three letter words.
So now we want to make words out of these three letters. And so this is one word, AUG, CUG, ACU, ACG. And you can see up here, they still have the three letter words, but at that time we really didn't care.
All we want to do is make a copy of it. Now what we're concerned with is what do these three-letter words mean, okay? And so these three-letter words mean different things.
And so AUG means methionine. CUG means leucine. ACU means therionine. And ACG also means therionine, okay? And so even though these are two of the same amino acids, they have two different words.
And again, that's the redundancy of the code. And so knowing what these three-letter words are will tell you what amino acid goes there. And this is the case for...
bacteria, you're talking about viruses, you're talking about plants, talking about animals are always the same. And so that's really a nice thing is that this thing, you can use this, whether you're talking about bacteria or you're talking about humans, talking about dogs, cats, anything in this case, it all means the same thing. And that's a really nice thing that you don't have to change it from organism to organism. It makes it a lot easier to understand.
Okay. So when we talk about transclavage, translation, what we're talking about is using those three letter words and building the protein code. Okay.
So typically we start with the start codon. And so that start codon means AUG. AUG is the three letter word that starts the process.
So whenever you have an AUG, that's typically what we call the start site. That tells you where the ribosome is supposed to bind on and the process is supposed to begin. That's where the first tRNA will bind and start building the protein.
This codes for AUG, codes for methionine. So methionine is typically the first amino acid that you have in all proteins because you have the AUG, which is the start. Okay.
What happens during this process is that the ribosome will bind on. The tRNA will come on to that, and it has a code that matches this code down here. Now, it's the same but different, and I'll talk about that in a minute. So what these three letter words stand for are what we call codons. On the tRNA, you have what is called the anticodon, which is just the opposite.
And so we'll see that in just a minute. But essentially, you have these different sites. That's where the tRNAs combine.
And based on where these things combine to, it brings the amino acid in, you build the proteins, and then along goes the process. And we'll take a look at all these processes here in just a minute. But this just kind of gives you an idea of how these things all fit together.
Okay. So here's initiation. So take a look at this video and this will give you an idea of how this gets initiated. Translation initiation is a process in which mRNA, initiator tRNA, and small and large ribosomal subunits associate with each other to form a complex.
Initiation of translation in bacteria begins when the 30S or small ribosomal subunit binds to the mRNA near its 5'end. This process is facilitated by a Scheindel-Garno sequence in the mRNA, which is complementary to a component of the small ribosomal subunit called 16S ribosomal RNA. Though a shorter sequence is shown here, the Scheindel-Garno sequence is actually 9 nucleotides long.
Initiation factor 3 also facilitates the binding of the mRNA to the small ribosomal subunit. Once the 30S subunit IF3 mRNA complex has formed, initiation factor 2 binds to the complex and promotes the binding of the initiator transfer RNA for N-formylmethionine to the complex. IF2 and IF3 Hydrolyze GTP as an energy source to promote the association of mRNA, tRNA, and the ribosomal subunits.
Translation initiation is completed when the large ribosomal subunit binds and IF2 and IF3 are released. Later, when translation is completed, IF1 is needed for the dissociation of the complex. Okay, so this goes into a lot more detail than... I really want you to know.
And essentially, this is just the initiation. So this is your AUG start site. And what happens is, is that it lines it up.
And this just gets into more detail of how it lines it up and everything else. And again, more detail than I want you to know. I just want you to know AUG is the start. You get the tRNA that comes in first. It binds in.
Then you get this 50S subunit that kind of clamps down on the top. And so when you get this clamping, this signals, okay, it's time to go. And then it starts the process along.
But again, it starts at this AUG site and that typically brings in methionine. And so we'll look at those things again as we go along. Okay. So again, once the initiation begins, you get a second tRNA with a complementary anticodon. And the anticodon, again, is just the opposite of the codon.
So let's take a look at the codon here. You have the AUG. AUG is the codon. You can see the anticodon is just the opposite. A's go with U's, U's go with A's, G's go with C's.
Okay, so here's the codon for the second tRNA. If we look at it, what is the anticodon? C goes with G, U goes with A, G goes with C.
So let's figure out this one here. What is going to be the anticodon for this one? If you have C, C, G, that's the codon. What would be the anticodon? I'll give you a second.
Okay, if you said G, G, C, you've got it. If you didn't get it, remember, C's go with G's. So in this case, the opposite is going to be G, the opposite is going to be G, and the opposite here is a G is going to be a C. Let's try this one, GCU.
You said CGA, you got it again. So this will be CGA because G goes with C, C goes with G, U goes with A. How about AUC?
Try that one. We'll finish up here. you said U, A, G, you'd be correct. So again, A goes with U, U goes with A, C goes with G. And so it's just knowing the opposite.
And remember in the case with RNA, there's no T's. So A's always go with U's and G's go with C's. And if you know the opposite, you always know the anticodon.
Okay. I'd like to ask that question on the test to see if you're paying attention. So be able, make sure you're able to do both the know how to understand the codon, and then know what the anticodon is. So the codon is the three letter word, and then the anticodon is just the opposite. Okay.
So practice those, write those out. And if you want me to give you some examples and that stuff, I'm happy to give you just some examples on the board and see if you got it. Okay. But practice, practice, practice. I like to ask those questions on the test just so that you understand the process.
Okay. All right. So once this happens, you have the tyranny that comes in.
These two amino acids will form a peptide bond. And now the process begins of how to build the protein. And we call this elongation because what you're doing is elongating the protein.
So as this continues on, you're going to elongate this protein. And that's called the elongation. Okay. And see the tRNA leaves.
And now you have the next one. Everything slides down. And once it slides down, it can fit into the next one. You form the next amino acid. Here it forms the peptide bond.
And then finally this will leave, this will keep it, and then this will slide down to the next codon to bring in the next tRNA. And then again, I have a video here just to show you how this all works. And this is called elongation. Chain elongation begins with the binding of a tRNA, which recognizes the next codon in the mRNA to the A site of the ribosome.
This is catalyzed by the EFTU transcription factor. and requires the hydrolysis of a GTP. Once the tRNA binds in the A site of the ribosome, the polypeptide chain is moved from the tRNA in the P site to the amino acid attached to the tRNA in the A site.
Peptidal transferase, a protein RNA complex present in the 50S ribosomal subunit, catalyzes the formation of this new peptide bond between the amino acids. The ribosome then translocates to the next codon. This process is promoted by elongation factor G and requires another GTP. This places the empty tRNA molecule in the E site of the ribosome. and moves the tRNA containing the growing polypeptide chain in the P site.
The next codon in the mRNA chain is positioned in the A site. The uncharged or empty tRNA in the E site then leaves the ribosome and a cycle of chain elongation is completed. Through subsequent cycles of chain elongation, the polypeptide chain continues to elongate one amino acid at a time.
Okay. So this just shows you it again, and this process continues, it's called the elongation. So you're elongating the protein and it continues to elongate as it goes on. And so that's the idea here. And so this will bring in the next here and a, this will connect, we'll slide over the next one, so on and so forth.
And that's the process that goes on with that. Okay. So translation continues until you reach a stop codon. Now there are four, there are actually three codons that don't have a tRNA.
And so here is the stop. codon, this UAG, that stop codon will say there's no tRNA and will cause the whole process to fall apart. And so we'll see how this goes here in just a minute.
We'll talk about termination and this is how you stop translation. Okay. So again, here are the, uh, termination codons, UAA, UAG, and UGA. These are the termination codons. There's no anti-codons for these guys.
What happens is that this says stop, and then the whole thing falls apart. And so what you'll see here in the next video is that it basically shows you what happens. It goes through, the whole thing falls apart, and voila, you're on to the next step. Okay, so let's see termination.
Termination begins when a stop codon appears in the A-site. Since there is no tRNA corresponding to the stop codon, a release factor binds in the A-site. The binding of the release factor causes the polypeptide chain to be cleaved from the tRNA.
The polypeptide is released and then the tRNA is released. In the last step, the two ribosomal subunits and the mRNA dissociate from each other. This completes the termination process. Okay, so again the process is over.
Everything basically falls off the mRNA. Now there could be another ribosome coming down the line and then we have these things called poly ribosomes where they're constantly making proteins and that but once it reaches the stop site that basically says stop here and fall off and so that's what happens in this process is that you reach the stop sign and then basically it would be coming to a stop sign and your car falling apart or not starting again and that's basically what happens here is that it just all falls apart and then you got to start over back at the beginning again all right it's termination okay so what is the trna is a molecule what does it do okay So does it contribute to the structure of the ribosomes? Does it adapt the genetic code to the protein structure?
Does it transfer the DNA code to the mRNA? Does it provide the master code for the amino acids? What does the tRNA do?
Now, it's not written very well here, so I'd like to change it, but I can't. So what do you think that this does? Okay, if you said B adapts the genetic code to the protein structure, I'd give that to you.
in that sense because really what it is doing is bringing the amino acid to the mRNA. It really doesn't transfer the DNA code to the mRNA and it doesn't provide a master code for amino acids in that sense because that's essentially the mRNA. I think some people could argue for D as well, but I kind of like B in this case.
So if you answered D or D, I'd give it to you. I think it's poorly worded. I would say which molecule is responsible for bringing amino acids to the ribosome. and you'd say the transfer RNA or which molecule has the anticodon and that would also be the tRNA.
Okay and like I said a lot of times you have these polyribosome processes where these things can happen over and over again so you can see that once transcription can be continuing on you have ribosomes as soon as they see the mRNA can start binding to this and this is a unique process in prokaryotes because the mRNA doesn't have to leave the nucleus because there is no nucleus in bacteria. So you can begin this process almost instantaneously. As soon as the mRNA is made, ribosomes bind on, and you can start making proteins.
And you can see that there is a multiple structure where you can have many, many different ribosomes binding to the same mRNA at once. And they can then make lots and lots of proteins. So this is a quick way for your cells and also the bacteria cells to make lots and lots of protein. And here you can actually see this process under an electron microscope.
So this does happen. We do know that it actually happens. Okay, we call these polyribosomes. Okay, so the last thing we're going to talk about is what is the difference between eukaryotes and prokaryotes.
Obviously, one of the big differences is eukaryotes have the nucleus. And so one of the big things that has to happen is that the RNA has to leave the nucleus before you can start translation. Prokaryotes, this doesn't happen, and that's because there is no nucleus. And so...
Again, a couple of differences is that it doesn't start simultaneously. You have to finish transcription first before you can start translation because you have to leave the nucleus in order for translation to take place in the cytoplasm. Another thing is that the AUG doesn't mean formal methionine. It's still methionine, but it's not formal methionine that you see in bacteria.
Again, in eukaryotes, one mRNA encodes for just one protein, whereas in bacteria, we're going to learn in the next section. that bacteria have operons where they have actually multiple genes on one piece of DNA. And so they can make multiple genes on one mRNA and make multiple things at once. Whereas in eukaryotes, it's one gene, one mRNA, one protein type of thing.
Okay. And then the other thing about eukaryotes is there's sections of DNA that have to be removed before you can actually start the process of translation. And these are called introns. And so in bacteria, we don't worry about it because they don't have introns. It's only found in eukaryotes that you have to splice the DNA.
And again, this has to happen before the RNA can leave the nucleus. And so the RNA has to be spliced before it can leave, before translation can begin. In bacteria, this can happen right away. There's no splicing that happens.
And so you have simultaneous transcription translation that goes on because no nucleus doesn't have to leave and no introns, which makes it really easy for these to happen. So again, this allows for... DNA to make proteins very quickly or for bacteria to make proteins very, very quickly.
Okay. And again, this just shows you the introns. So this is the, the mRNA, you have these areas of DNA that are RNA that are not coded. And so you have to take these out.
This is found in eukaryotes. And so I just bring this up because again, you may have learned this already and said, Oh, what about splicing all these other things? We don't worry about it in bacteria because bacteria don't have to worry about.
strands are getting rid of introns. There are no introns in bacterial DNA. Okay.
All right. And this will just show you, again, this video showing you the difference between prokaryotes and eukaryotes. And I think it gives you a good example.
And then once we get this, we're done for today. Okay. Gene information is processed differently in prokaryotes and eukaryotes.
Because prokaryotes lack a nuclear envelope, messenger RNA, mRNA. can associate with ribosomes in the cytoplasm as the mRNA is being formed. Translation of mRNA into protein begins before transcription is complete. Individual bacterial mRNA molecules often contain transcripts of several genes.
By placing genes with related functions on the same mRNA, bacteria coordinate the synthesis of these proteins. These clustered genes are referred to as an operon. Because eukaryotic cells possess a nucleus, their mRNA must be completely formed and must pass across the nuclear envelope before translation. Eukaryotic mRNAs are modified before they are translated.
Introns are removed and the remaining exons spliced together. A 5'cap and a 3'poly-A tail are added. The processed mRNA travels to the cytoplasm where translation occurs. In contrast to prokaryotic mRNA, eukaryotic mRNA usually specifies only a single protein. Okay, so the biggest thing that you get out of this is that in eukaryotes, the process is much more complex.
You have to splice, you have to add these caps, and then you have to leave the nucleus before translation can begin. Bacteria, you don't even have to worry about that. That's all can take place.
simultaneously. Basically you have transcription and then translation can happen just after transcription. So that's really what I want you to get out of it is that there's a lot less processing that takes place in bacteria can happen simultaneously. And the other big thing is that you can have multiple genes on one mRNA. Eukaryotes, there's only one gene, one mRNA, one protein.
Okay. And so that's how we look at it that way. All right. So we've made it to the end. What we talked about today first is microbial genetics.
And again, this is a study of genes and bacteria and other various microbes. Again, bacteria have a single circular piece of DNA, roughly four to five million base pairs. They also contain plasmids, which are included in the genome. And these are very small pieces of DNA, anywhere from 1,000 to 10,000 base pairs.
And again, Only one letter, one gene per chromosome. And so, again, the phenotype. So we really don't worry about genotypes and phenotypes in the sense that doing punnett squares and dominance recessive, we only care about what do the bacteria look like and do they have the genes.
Okay, remember for the DNA structure, we talked about the nucleotide. The nucleotide is the phosphate, the sugar, and the base. And remember, in DNA, they run in opposite directions, 5'', 3'', and 1''.
three prime five prime in the other direction again opposite and that a's and t's bind together and g's and c's bind together in the dna okay we get into replication replication involves just copying dna no rna involved so this is just making a new strand and this is only when you want to divide your cells okay it's semi-conservative you open up the dna with helicase and you make two new strands so you have the two old strands two new strands get made on the inside okay We talked about leading and lagging strands. The leading strand is the one complete piece of DNA that's made 5'to 3'. The lagging strand are those small little segments that have to be glued back together. And in bacteria, again, it's a circle.
So you start at the origin of replication and you work your way around to the other side. And so it opens up to form two new pieces of DNA. Okay, and then we talked about transcription and translation.
These are on all the time because you always need to make proteins. And again, It's two steps in this process. This is how you make proteins. First step is transcripting, which is making a copy of RNA.
So what you're doing is not making a copy of DNA. You're making a DNA or RNA copy of DNA. And again, there's a slight difference in RNA.
RNA has ribose and has uracil. So no T's in RNA. And what you're doing is reading that top strand. So you would, or you're reading the bottom strand and you're making a copy of the top strand.
So when you make an mRNA, When I give you the five prime to three prime sequence, you just basically rewrite that sequence and just replace the T's with U's. We'll keep it easy. Okay. The bottom strand is red.
And then once the RNA is made in prokaryotes, it can be translated right away. Translation is now taking all those individual letters and making three letter words. Those three letter words are called codons.
Those codons tell you what amino acids are going to be brought in. Again, the tRNA, which is responsible for bringing the amino acid to the ribosome, brings it in, has the anticodon. That anticodon recognizes the codon.
And so, again, remember, it's just the opposite. So if I have AUG as my codon, the anticodon is going to be UAC. Okay.
And so that's just the opposite. So AUG, UAC. Okay.
Just think of the opposite for the anticodon. And again, three-letter words. It goes through the process of initiation. It always starts at AUG. You have the elongation where you add more tRNAs to bring more amino acids in as it chugs down the line.
And then finally, you come to the stop codon where the stop is. Again, the process falls off at that point in time and the protein disassociates along with the ribosome. And then now you are ready to begin again.
Again, a couple of differences between the eukaryotes and prokaryotes. Eukaryotes have to splice and do all these things. You got to go through the nucleus.
For prokaryotes, you don't have to worry about any of that stuff. There's no introns. All takes place in the cytoplasm. And so you can basically begin this process simultaneously.
So it's a little bit more efficient in the prokaryotes. Okay, so we've come to the end. If you have any questions, please feel free to ask me in class or via email. Happy to answer your questions.
I'm glad that you're watching these videos and I'll see you next time.