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Current time:0:00Total duration:8:57

Hybridization (microarray)

Video transcript

so in this video we're going to be talking about something known as DNA hybridization Tignes hybridize alright so in this video we're going to be talking about something known as DNA hybridization DNA hybridization now what is DNA hybridization well basically what it so let's work through an example to try and explain what DNA hybridization is so let's imagine that we have two cells so over here we have cell a and over here we have cell B now let's imagine that cell a is a cancer cell so this is a cancer cell and cell B over here is a normal cell this is normal now cancer cells basically have the ability to proliferate and grow and grow and metastasize and move throughout the body so basically they have this unregulated cell growth and the reason that the cell growth is unregulated is because there are various mutations that cause changes in the proteins that are expressed and changes in the regulation of the cell cycle and there are hundreds and hundreds of different mutations and hundreds of different proteins that could be affected and all of them can lead to cancer now what is producing different proteins and different amounts now what are kind of the two options that we have for certain genes so let's imagine that we have gene a over here so if this is gene a what are the two options either gene a can be upregulated or it can be down regulated so if it's up regulated then what we have is we have the gene products which is mrna and eventually protein we have a lot more of the mRNA and the protein that gene a encodes for and what that basically means is that let's imagine that gene a encodes for a protein that will induce cellular proliferation and will allow that cell to go in metastasize throughout the body well if we have a lot more of gene a being Express either because the promoter is upregulated or for whatever reason now we have lots and lots of this protein that basically allows cellular proliferation to occur we have lots of this protein floating around the cell and we have this cancer cell proliferation uncontrollably so another option is if we have gene B so if we have gene B gene B could be down regulated and that basically means that the gene B isn't producing its gene product and what if that gene product were something that basically stopped this cell from polar frating well if we have less inhibition then we basically have more proliferation and the third option for any specific gene in a cancer cell so let's say gene C is that there's no change so there's just no change so what we want to do is use DNA hybridization technology in order to assay the gene transcription profiles of a cancer cell compared to a normal cell and in order to do that we need something we need to use something known as a micro array so a micro array now what is a microarray well array basically means that we're asking a whole bunch of different things and in this case we're asking the transcription profiles of a bunch of different genes and micro just means that it's small so this could be as small as a chip so let's imagine that we have a microarray chip so let's say that we've got this chip and it's basically just this this Square and this chip has a lot of different holes in it so let's imagine that we've got just lots and lots of holes so we have just hundreds of these holes and I'll just drop you for simplicity sake so we have a bunch of these holes on the microarray chip now these holes are actually little tiny wells they're microscopic wells so if we actually looked at this from the side sewed so here's the chip we're looking at it from the top so it's lying on the table we're just looking down at it if we looked at it from the side one of these wells would look like this and inside the well would be the would be a mRNA complementary mRNA strand so we've got just lots and lots of these little complementary mRNA strands and what are they complementary to well they're complementary to a specific gene so let's say that inside one of these wells let's draw another well over here let's say that inside one of these wells we have the complementary mRNA to gene a so we've got the complimentary mRNA to gene a now let's imagine that in this cancer cell gene a is upregulated for whatever reason and if gene a is upregulated it's being over transcribed and that means that there's lots and lots of the gene a mRNA floating around from the cell so there's just a bunch of the gene a mRNA and this is in comparison to the normal amount of gene a products which might just be a few gene a mRNA s now what we can do is we can take this cell and we can break it apart and we can label the mRNA with a certain color so let's say that I label each one of these mRNAs with a yellow fluorescent label so let's imagine that I labeled every single one of these mRNAs with a yellow fluorescent label and let's imagine that I labeled the mRNA in the normal cell with a blue fluorescent label now what I can do is I can break these cells apart and I can basically add the intracellular contents to this well so I can add it to this well and since I have lots and lots of this mRNA that's labeled yellow what I'm going to have I'm going to have a lot of the mRNAs binding to the complementary strands and so I'm going to have a really bright yellow well and when I add the normal cell intracellular contents I'm going to have some blue so I'm going to have lots of yellow and a little bit of blue and what that will basically look like is it'll really just you won't be able to see the blue it'll really look like just a bright yellow dot so let's imagine that this is the well it'll look like a bright yellow dot and a computer can scan every single one of these wells and basically decide okay is it a brighter yellow or is it a brighter blue if it's a brighter yellow color if you see mainly yellow that means that you have a lot more of that specific genes products being expressed in the cancer cell compared to them normal cell now let's imagine that we look at a downregulated gene so if we look at it down regulated gene let's just draw another well so let's draw well over here now if we look at a down regulated gene we've got lots of the gene via mRNA two complementary strands inside this well and we're going to have very few of the gene be mRNA in the cancer cell and then a lot more in the normal cell and once again we'll label the gene be mRNA with a yellow fluorescent label and over here again I'm sorry over there we're going to label it blue so we'll label it with this blue fluorescent label and once again we're going to lyse the cells and we're going to expose the intracellular contents to this well and what we're going to have is we're going to have very few of the gene b products binding and we're going to have a lot of the normal cell of the gene B byproducts in the normal cell binding so when you look at this well it's going to pop up as mainly blue and once again the computer is going to read this and it's going to notice oh well this well has mainly a blue fluorescent label which means that the normal cell is expressing a normal amount and there's a lot less of that gene being expressed in the cancer cell so this is kind of the idea of a microarray chip and assing the gene expression profile in a cancer cell versus a normal cell it's able to tell you whether specific gene is upregulated or downregulated and you're also able to see if a specific gene has no change and if there's no change then instead of seeing either a yellow or a blue dot you would see something kind of in the middle so maybe you'd see a green dot and that's basically a quick way in order to look at a whole bunch of different genes on a single chip and try and quickly determine which gene is upregulated or downregulated in the cancer cell compared to a normal cell and this can help you tailor your therapy so let's say that you know that this that this well right here is for a specific protein and you have a drug that's able to target that protein well now you're able to tailor your therapy for that individual patient use this microarray technology