- DNA technology questions
- Gel electrophoresis
- Polymerase chain reaction (PCR)
- DNA libraries & generating cDNA
- DNA cloning and recombinant DNA
- Hybridization (microarray)
- Expressing cloned genes
- Southern blot
- DNA sequencing
- Gene expression and function
- Applications of DNA technologies
- Safety and ethics of DNA technologies
Visit us (http://www.khanacademy.org/science/healthcare-and-medicine) for health and medicine content or (http://www.khanacademy.org/test-prep/mcat) for MCAT related content. These videos do not provide medical advice and are for informational purposes only. The videos are not intended to be a substitute for professional medical advice, diagnosis or treatment. Always seek the advice of a qualified health provider with any questions you may have regarding a medical condition. Never disregard professional medical advice or delay in seeking it because of something you have read or seen in any Khan Academy video. Created by Ronald Sahyouni.
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- Did anyone else think this video didn't flow correctly as understanding the content?(88 votes)
- Yeah, the title suggests the focus is on hybridization, but the video focuses on microarray and only seems to mention hybridization in passing. I honestly went through the entire video wondering if it had been mislabled somehow.
I think the point is that the microarray uses hybridization (the complimentary strands in the well and the ones being examined).(3 votes)
- So what is DNA hybridization?(19 votes)
- DNA hybridization is a method used to compare the genomic similarities between different DNA strands. The procedure involves denaturing double stranded DNA (dsDNA) from two different samples, and then allowing the single strands to re-anneal into dsDNA such that one strand is from one sample and the other strand is from the other sample, producing a "hybrid" dsDNA. The stronger the hybrid strands are able to hydrogen bond together, the more similar the original DNA samples were to each other (stronger binding = more complementary base pairs = more similar sequences).
Really, this video is more about microarrays than the process of DNA hybridization itself.(25 votes)
- Are both disease and normal fluorescent labelled mRNA put into the the same well on the microarray??(5 votes)
- All of the wells in the micro-array chip are filled with the intracellular contents of both the cancer and normal cells. Each well has a unique set of complementary-mRNAs (c-mRNAs) to detect a particular gene. Only mRNAs (from each cell's intracellular fluid) which match a given well's c-mRNA will anneal or "stick" to that well. Because the cancer cell mRNAs are labeled with a different colored fluorescent marker than the normal cell mRNAs, this method allows you to tell which genes occur more frequently in each type of cell.(14 votes)
- This link actually says we are using dna strands in the microarray chips?
But in the video he says we are using RNAs in the chip ?(4 votes)
- The mRNA can be reversed transcribed into complementary DNA (cDNA) using the enzyme reverse transcriptase. This makes analysis easier because DNA is much more stable than RNA, which is prone to degradation. Note that cDNA is NOT the same as genomic DNA (gDNA), located in the nucleus of cells. Analysis carried out using cDNA still assesses gene expression levels.
- Whats is actually put in the CHIP well?
mRNA or cDNA?(1 vote)
- The well is lined with complementary RNA to the mRNA sequence of the gene being studied. That way, the mRNA with the 'glow sticks' from the cells will stick to the well (hybridizing with the complementary RNA) when injected. In the example, he didn't use DNA. Hope this helps!(7 votes)
- This video concentrates too much on microarray, you only need to know about hybridization. premedhq.com summarizes this concept as:
**Denaturation/annealing of DNA
-These two techniques are widely used in PCR (polymerase chain reaction)
-Denaturation: double stranded DNA comes apart due to heating or a change in pH (increase in temperature in the environment)
-Annealing: If the temperature returns to normal (or the environment returns to normal), two single strands form double stranded DNA again due to the complementary nucleotide sequences and to random molecular motion
-This is a much slower process compared to denaturation
-Requires salt to neutralize the repulsion of sugar phosphate backbones from each strand
-Molecular biology technique that compares and analyzes the degree of genetic similarity between identical or related DNA sequences
-Measures the genetic distance between two species
-Utilizes the denaturation of two different DNA sequences, then anneals single stranded DNA from each(3 votes)
- At3:09you mention that you want to assay the gene transcription profiles of the cancer cell. What does "assay" mean?(3 votes)
- Shouldn't the well contain cDNA complementary to the mRNAs? Because the speaker keeps calling the contents of the wells mRNA... Or am I just confusing something?(2 votes)
- How do you know what complementary mRNA strand each well would have if you're going to label all the genes with the same 2 colors?(2 votes)
- [Voiceover] So in this video, we're going to be talking about something known as DNA hybridization. DNA hybridization. Alright, so in this video we're going to be talking about something known as DNA hybridization. So, 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. So, 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 effected. And, all of them can lead to cancer. Now, one is producing different proteins in 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 downregulated. So if it's upregulated 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 and metastasize throughout the body. Well, if we have a lot more of Gene A being expressed, 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 proliferating uncontrollably. So, another option is if we have Gene B, so if we have Gene B. Gene B could be downregulated. And that basically means that Gene B isn't producing its gene product. And what if that gene product were something that basically stopped this cell from proliferating. 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, we 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 to use something known as a microarray. So, a microarray. Now, what is a microarray? Well, array basically means that we're assaying a whole bunch of different things. And in this case, we're assaying 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 square. And this chip has a lot of different holes in it. So let's imagine that we've got lots and lots of holes. So we have just hundreds of these holes. And I'll just draw a few for simplicity's sake. So we have a bunch of these holes on the mircoarray chip. Now these holes are actually little tiny wells, they're microscopic wells. So if we actually looked at this from the side, so here's the chip, we're looking at it from the top. It's lying on the table, we're 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 complimentary mRNA strand. So, we've got just lots and lots of these little complimentary mRNA strands. And what are they complimentary to? Well, they're complimentary 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 complimentary 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 overtranscribed and that means that there's lots and lots of the Gene A mRNA floating around in this 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 mRNAs. 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 the 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'll 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 gene's products being expressed in the cancer cell compared to the normal cell. Now, let's imagine that we look at a downregulated gene. So it we look at a downregulated gene, let's just draw another well. So let's draw a well over here. Now, if we look at a downregulated gene, we've got lots of the Gene B mRNA, the complementary strands, inside this well. And we're going to have very few of the Gene B mRNA in the cancer cell and then a lot more in the normal cell. And once again, we'll label the Gene B 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 lice the cells and expose the intracellular contents to this well. And what we're going to have, 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's 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 assaying the gene expression profile in a cancer cell versus a normal cell. It's able to tell you whether a 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 a cancer cell compared to a normal cell. And this can help you tailor your therapies. So, let's say that you know 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 using this microarray technology.