this last section on chapter 8 is on mutations and mutations are going to be changes in the DNA we usually think about mutations as being mistakes I often call mutations The Good The Bad and The indifferent sometimes they're good so that new alals arise so if let's say there was a mutation and a bacteria now has the ability to be resist to an antibiotic it can live in the presence that's good for the bacteria bad news for us but that's a good mutation for the bacteria sometimes be bad so they can cause the proteins not to function and sometimes they can be indifferent where there is a change in genotype and the genotype is the DNA but it doesn't change the phenotype or the outcome and in that case even though the DNA was changed it doesn't really make a difference so often time with mutations we talk about genotypes and phenotypes genotypes are what are determined by the DNA so if there is a mutation we can change the genotype mutations always change the genotype but they may or may not change the phenotype the phenotype is going to be the physical characteristic of an organism as we saw with proteins our proteins are made up of amino acids and the sequence of amino acid is going to be very important for that protein to fold and for it to function if there was a change in genotype and we ended up with a different codon if you remember that Cod on table we could have four codons code for an amino acid so if we ended up with a mutation that changed the codon but we still ended up with the same amino acid we didn't actually change the phenotype and in that case the mutation didn't really make a difference so in this section we'll talk about the different types of mutations as well as how we can also study them in bacteria so we will look at different types of mutations like a duplication this is when one nucleotide is repeated a deletion is when a nucle tide is removed an inversion is when the segment of the sequence is kind of flipped around and insertion or translocation this is when we have a segment of DNA maybe further down the line and it's just kind of plunked right in the middle of DNA breakage is easy to think about because part of it is just lost and we're going to talk a little bit more in depth about a frame shift mutation this is when we could have an insertion or or deletion of a single nucleotide but because of how the codons are red it actually shifts the frame that the codons are red and so we could end up with completely different amino acids so this picture right here is showing the DNA and a point mutation a point mutation would be a change in the nucleotide so if you took a look at picture a so we've got the original DNA and the code is a AA and so on our messenger RNA transcript our codon is uuu that codon will code for the amino acid phenyalanine so that is what the amino acid we need in that particular order if you take a look at picture B that third a is changed to a t so we have a base substitution so now our codon becomes uua this codes for lisine so we're not getting pheny alanine is that a big deal it could be it could affect how that polypeptide folds if you see in the third case in C that third a is changed to a g so now our codon is uu and we're getting phenol alanine so even though we changed the genotype we did not change the phenotype so that that's what those extra codons kind of give us as that little insurance policy so we can change the genotype sometimes it can actually change the phenotype sometimes it doesn't in this case of that third base substitution so I think it's a little easier to actually think about letters and words rather than um a code so I've got the original DNA code but I'm using letters so up at the very top a b CDE e that is our original DNA code so if we had a duplication and we duplicated that b now our code is a BB CDE if we have a deletion where we remove the B now our code is a CDE and if we have sort of an inversion where we flip around BCD now our code in green as you see becomes a dcbe a translocation or insertion this is when further down the line we just kind of take that code of DNA and kind of plunk it right in the middle so further down in the alphabet we've got R and S so as you see with this translocation or this insertion our code now becomes AB RS CDE the last one is a little easier to kind of wrap our head around this is a deletion so we just kind of take off d& and that breakage and now we have that ABC as our code I told you we were going to talk a little bit more about frame shift mutations and so I think it's a little easier to see with words and a sentence so this is a a silly sentence the dog ate the cat but we've got three letter words and those three letters represent our codons now this is a silly sentence but we can at least recognize those words so we know the dog ate the cat we can recognize that if we have a simple deletion in the case of a frame shift mutation this is just taking one little change in this case we're going to delete the E we could have easily inserted another nucleotide another letter but in this case we're going to delete that at e so with that frame shift mutation we are shifting how we're reading the frame of those letters in the case of the protein we would be shifting how we read those codons so when we squish these letters together our sentence the dog at the cat now becomes Theta AA T hack at these are nonsense words they make no sense in terms of the protein and getting different codons we can actually get different amino acids this can have a big consequence on our proteins so if we took a look at an actual example if you take a look at the normal DNA code and codons so up top we've got our DNA code and we're getting codons these codons are going to get us the sequence of amino acids phenol alanine Serene histadine l and lysine so in that second block now what we're going to do is we're going to remove that guanine so we're going to take out that nucleotide this is a simple deletion so now we're going to squish all of those letters together and we're going to get different codons so we're on track with the first one with phenol alanine but instead of serene we're getting Arginine instead of histadine we're getting thine instead of lisine we're getting a stop C on this is a big problem so if this polypeptide was supposed to be thousands of amino acids long we only have three amino acids and we end up with a stop Caton so that is what we would call a loss of function mutation that's bad no way can that polypeptide fold no way can that protein function if we take a look at the third example now we've got a simple insertion now we're in inserting ayine after that guanine again we're going to push them together so now we're going to get different codons we're still on track with phenol alanine instead of serene we're getting veine instead of histadine we're getting alanine we're back on track with lisine instead of Lysine we're getting glutamic acid and so forth is that a big deal it could be because remember those amino acids differ a little bit chemically so as they're sitting in line together that polypeptide may not fold as it's supposed to with that protein and that protein may not function so even though we have a very small change the way that we shift that frame can actually have big consequences we can actually study mutations in bacteria so oxrs are bacteria that have acquired mutations that affect their ility to make and use certain nutrients so we can actually use this to our advantage when we're studying mutations uh a normal unmutated bacteria would would be what we would call a protot tro I'm going to talk lastly about a little experiment where we can use an Oxo trro to determine whether or not a chemical or drug that we're testing is a mutagen we know that mutations can spontaneously occur so there can just be mistakes that happen in the DNA in DNA replication we talked about DNA polymerase and that builds DNA DNA polymerase has a little proofreading capability where it can kind of change out a nucleotide if it isn't matching but only to a certain extent so mutations can spontaneously happen we also know that mutations can be induced by mutagens so maybe certain chemicals or radiation they can cause mutation so we know that different chemicals and things can also cause mutations not only in bacteria uh but in humans as well one of the things that we'll talk about when we get into viruses is that sometimes viruses can cause mutations um and we know that viruses there are some viruses that can cause cancer so here's the thing about bacteria is that if they have DNA damage if they have mutations and often times times if they're exposed to radiation these thyine dimers can form where we have these two thines that are kind of stuck together bacteria can be very smart and sneaky and that they can actually repair this damage and so they can actually take these enzymes kind of like these molecular scissors that can actually cut that dier and then remove them so they can actually repair some of their damage to a a certain extent that's good news for the bacteria if they have mutations there were a couple of scientists Lura Del Brook that wanted to determine how mutations that um allowed resistance to antibiotics arose and so what they did was they designed a fluctuation test to actually test these two hypotheses so hypothesis one was that if mutations that cause resistance to antibiotics happen spontaneously then when they are growing in the absence of the back of the antibiotic there's going to be a great fluctuation in the number of bacteria that have that resistance mutation so this is just to say mutations happen spontaneously doesn't matter if they're exposed to that antibiotic or not they're just going to happen the second hypothesis they had said these mutations for resistance can actually arise only in the presence of the antibiotic so that bacteria had to be exposed to that antibiotic in order to have that mutation arise so they did a test this is what is called the fluctuation test and we're not going to get too bogged down on the weeds in the detail with this but what they did was they grew bacteria in individual containers and then in a big flask the take-home message from this fluctuation test and the results they found proved hypothesis one that mutations just happen spontaneously that bacteria don't need to be in the presence of that antibiotic to get those resistance genes from mutations that they just happen spontaneously so this is what Loria Del Brook found in their fluctuation test the last test that I'm going to talk about is the ases test and the Aims Test this is actually a Toxicology test that determines whether or not chemicals drugs whatever it is that a company is is trying to put out as a product is a mutagen and causes mutations or can cause cancer this is a very common toxicological test um and if you were to actually buy a chemical from a supply house it would actually come with a material data sheet on that material data sheet it would talk about the chemical if it's flammable and usually the last thing that they talk about is this Aims Test whether or not this chemical causes mutations this is going to be important if we're working with chemicals in the lab because we want to take some safety precautions if a company a pharmaceutical company is putting out a drug they want to prove that this drug does not cause mutations does not cause cancer so with this ases test it actually uses an oxit trophic form of salmonella remember this oxit trophic form has special nutrient requirements so with this particular oxota it cannot um live without the supplement of histadine it actually has to have histadine this amino acid in its media because it cannot make it itself so in order to grow it you would need a supplemental media with histadine in order for it to grow so what we would do in this as test is that we would actually take this oxitop stanilla we would plate it on an augur plate that is lacking that histadine now because this salmonella would require that histadine in the media it would actually die on that plate and so this is how the scientists would test whether or not that chemical or drug they were testing formed mutations so they would take a uh filter paper disc soak it in the chemical that they wanted to test and then put that on the lawn of growth then they would incubate it if that chemical did not cause mutations then that bacteria would not be growing so we would not see growth if it did did cause mutations then we would see growth and that means that that particular oxota develop mutations to allow it to now make that histadine so that it could live so if we saw growth that's a good indication that that chemical caused mutations so these results of an aim test would be on that material data sheet so that we would know that we needed to take special precautions because that particular substance was known to cause mutations