Introduction to Polymerase Chain Reaction - Online Tutor, Practice Problems & Exam Prep

Polymerase chain reaction (PCR) is a rapid technique used to amplify specific DNA sequences in a test tube, differing from DNA cloning, which occurs in living cells. PCR requires key components: template DNA, two complementary primers, thermostable DNA polymerase (often Taq polymerase), and all four deoxyribonucleotides. The amplification process occurs in cycles, with the number of DNA copies calculated using the formula $2^n$, where n is the cycle number. This efficiency makes PCR invaluable in fields like forensic science.

Introduction to Polymerase Chain Reaction

In this video, we're going to begin our introduction to a technique that's called polymerase chain reaction. And so when a researcher studies the DNA sequence of a specific gene in a genome, they actually need to make many copies of that gene in order to study it. And so this technique, polymerase chain reaction, which is commonly abbreviated as just PCR, is a technique that's used to rapidly isolate and amplify a specific sequence of DNA. Now, the term amplify is just referring to the process of making more copies of that DNA. And so, this is kind of similar to DNA cloning in a way because DNA cloning is also going to make more copies of the DNA. However, unlike DNA cloning, which is going to be using living cells to make more copies of the DNA, polymerase chain reaction or PCR will take place in a test tube. And so, of course, the test tube is not going to include living cells. We’ll be able to talk more about this process of polymerase chain reaction and PCR as we continue to move forward in our course. But for now, this here concludes our brief introduction to polymerase chain reaction or PCR, and I'll see you all in our next video.

Why do we use PCR?

So now that we've briefly introduced polymerase chain reaction or PCR in our previous lesson video, it's fair to ask why we use PCR. Well, it turns out that scientists will use PCR for many different reasons. And that's because PCR is a quick and efficient process for generating many identical copies of DNA in a test tube. Now DNA cloning, which occurs inside of living cells and does not occur in test tubes, is going to be more accurate, but it's actually a less efficient process. And that's because, recall, DNA cloning occurs inside of cells, and although there are fewer mutations inside of cells, it's going to take a lot longer to amplify the DNA inside of cells. And that's because a lot of times cells take 24 hours or more to be able to grow, and then after you grow the cells, you have to isolate the DNA, and that's a whole another step within itself. And so PCR is much quicker and much more efficient. And so in our example, down below, we're going to take a look at, just in general, how PCR, or what PCR can do. And so, it says here that PCR can be used to amplify the amount of DNA taken from a crime scene so that a detective can actually investigate the DNA. And so down below over here, notice we have this scientist that has a test tube here with some DNA that perhaps was found at a crime scene, but notice that there's not a whole lot of DNA within this sample. And so notice the scientist is saying, "I wish I had more of this particular DNA of interest, so that he has enough DNA to be able to run tests on it." And so this is where PCR can come into play because the process of PCR, polymerase chain reaction, can amplify the amount of DNA to make many identical copies of the DNA as we see over here. And so the scientists are saying, "Wow, that amplified very quickly." And the process of PCR can be done in a relatively short period of time, maybe something like 2 hours, 1 and a half, 2 hours, whereas DNA cloning, again, is going to take much, much longer, which it could take well over 24 hours in many cases. And so, why do we use PCR? To make many identical copies of DNA in a test tube very quickly and efficiently. And so again, as we move forward in our course, we're going to continue to talk more and more about PCR and the steps of PCR. So I'll see you all in our next video.

PCR is used to _____.

• A. Amplify a single gene or small sequence of DNA.
• B. Create DNA without introns.
• C. Insert foreign DNA into a host cell where it can be replicated.
• D. Cut segments of DNA creating complimentary sticky ends.
• E. None of these.

Components of a Polymerase Chain Reaction

So before we can talk about the details of each of the steps in polymerase chain reaction or PCR, it's first helpful to talk about the components of Polymerase Chain Reaction. And so recall from our previous lesson video that PCR differs from DNA cloning in the location that the DNA is replicated. And so PCR occurs within a test tube, whereas cloning is going to occur within a cell. And so the components that go into the test tube during PCR include these components that we have listed down below. And so the components of the PCR mixture include a here is going to be a template DNA. And so the template DNA is going to contain the sequence of interest for the study that the scientist is interested in amplifying and creating more copies of it. And so down below in this table on the left-hand side, you can see that the components of the PCR mixture in this table correspond with the components of the PCR mixture that we have up above in the text. And so the template DNA, you can see, is right here in this image down below. Now for the second component, B, there are 2 primers that are required, 2 DNA primers. These 2 primers are going to be complementary to the opposite strands of DNA and are going to be oriented towards each other and serve as the starting point for the amplification. And so down below, what you can see is part b here are the DNA primers, which are gonna be small little DNA molecules, that are oriented opposite from one another. So notice that one goes from 5′ to 3′ left to right, and the other goes 5′ to 3′ right to left. And we'll talk more about these DNA primers and exactly how they're used as we move forward in our course. Now the third component that's going to be needed is a thermostable DNA polymerase. And the thermostable DNA polymerase is going to be the main enzyme that's used to synthesize the sequence of interest to amplify and make more copies of the DNA of interest. And so down below here in the 3rd component, c, you can see that we need a thermostable DNA polymerase, which is really, commonly going to be a DNA polymerase called Taq polymerase, which we'll get to talk more about Taq polymerase as we move forward in our course. Now the 4th and final component here that's going to be needed, in the PCR mixture is, d here, which is going to be all 4 deoxyribonucleotides or all 4 DNA nucleotides that are used to synthesize DNA. And so down below here in this image, you can see that d is going to include all 4 of those DNA nucleotides, which you can see over here, which includes these nucleotides that have the nitrogenous bases t, a, c, and g. And so these are the main components that are needed for a PCR mixture.

Now over here on the right-hand side, what we're showing you is a little glimpse of the process of amplifying a gene with PCR. And so, of course, you're gonna start off with your template DNA, which we have here in this first column. And it turns out that PCR, polymerase chain reaction, actually occurs in a series of cycles. And so, the first cycle you can see is going to amplify the DNA. The second cycle will amplify the DNA even more, and the third cycle will amplify the DNA even more. And continuous cycles will amplify the DNA more and more and more. And so there's actually a PCR formula that can be used to determine the number of new copies of template DNA that are made at each cycle. And the formula is relatively simple. It's just 2n where n is gonna act as an exponent here, and n is just a variable that represents the number of PCR cycles. And so for example, if we wanted to calculate the number of new copies of template DNA in the first cycle of PCR, all we need to do is take 21 since one is representing cycle number 1. And so 21 is equal to 2. And so notice that after the first cycle, there are 2 new copies of DNA. Now after the second cycle, if we wanted to calculate how many new copies of template DNA there would be, you just take 22. And 22 is equal to 4. And so you can see that there are 4 new copies of the DNA after cycle number 2 of PCR. And the same goes for cycle number 3. If we want to calculate the number of new copies of template DNA, you just take 23, which is the number of cycles, cycle 3. And 23 is equal to 8. And so you can see that there are 8 copies of the DNA, after cycle number 3. And so, we'll be able to get some practice applying these concepts that we've learned as we move forward in our course. And then later, we'll get to talk about the actual steps that are involved in polymerase chain reaction or PCR. So I'll see you all in our next video.

The polymerase chain reaction:

• A. Doubles the template sequence of DNA with each cycle.
• B. Makes a single copy of the template sequence of DNA with each cycle.
• C. Is used to determine the sequence of a region of DNA.
• D. All are correct.

Here’s what students ask on this topic:

What is Polymerase Chain Reaction (PCR) and how does it work?

Polymerase Chain Reaction (PCR) is a technique used to rapidly amplify specific DNA sequences in a test tube. The process involves several key components: template DNA, two complementary primers, thermostable DNA polymerase (often Taq polymerase), and all four deoxyribonucleotides. PCR works through a series of cycles, each consisting of three main steps: denaturation (heating the DNA to separate strands), annealing (cooling to allow primers to bind to the template), and extension (DNA polymerase synthesizes new DNA strands). The number of DNA copies doubles with each cycle, following the formula $2^n$, where $n$ is the cycle number.

Why is PCR preferred over DNA cloning for amplifying DNA?

PCR is preferred over DNA cloning for amplifying DNA because it is quicker and more efficient. While DNA cloning involves growing cells and isolating DNA, which can take over 24 hours, PCR can amplify DNA in just 1.5 to 2 hours. PCR occurs in a test tube without the need for living cells, making it a streamlined process. Although DNA cloning is more accurate with fewer mutations, PCR's speed and efficiency make it ideal for applications like forensic analysis, where rapid results are crucial.

What are the main components required for a PCR reaction?

The main components required for a PCR reaction include: 1) Template DNA, which contains the sequence of interest; 2) Two DNA primers, complementary to the opposite strands of the DNA and oriented towards each other; 3) Thermostable DNA polymerase, commonly Taq polymerase, which synthesizes new DNA strands; and 4) All four deoxyribonucleotides (dNTPs), which are the building blocks of DNA. These components are mixed in a test tube and subjected to cycles of heating and cooling to amplify the DNA.

How does the number of DNA copies change with each PCR cycle?

The number of DNA copies in a PCR reaction doubles with each cycle. This exponential amplification can be represented by the formula $2^n$, where $n$ is the number of cycles. For example, after the first cycle, there are 2 copies of DNA; after the second cycle, there are 4 copies; and after the third cycle, there are 8 copies. This rapid increase allows for the generation of millions of copies of the target DNA sequence in a relatively short period.

What is the role of Taq polymerase in PCR?

Taq polymerase is a thermostable DNA polymerase used in PCR to synthesize new DNA strands. It is derived from the bacterium Thermus aquaticus, which thrives in hot environments, making Taq polymerase stable at the high temperatures required for PCR. During the extension step of each PCR cycle, Taq polymerase adds nucleotides to the primers, creating new DNA strands complementary to the template DNA. Its ability to withstand repeated heating and cooling cycles is crucial for the efficiency and success of the PCR process.