Components of a Polymerase Chain Reaction

by Jason Amores Sumpter
201 views
1
Was this helpful ?
1
So before we can talk about the details of each of the steps in preliminaries chain Reaction or PCR, its first helpful to talk about the components of preliminaries, chain reaction. And so recall from our previous lesson video that PCR differs from DNA cloning and 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, uh, these components that we have listed down below. And so the components of the PCR mixture include, uh, a here is going to be a template d n a. And so the template d n A. 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, uh, corresponds with the components of the PCR mixture that we have up above in the text. And so the template D n A you can see is right here in this image down below. Now, for the second component, B uh, there are two primers that are required to DNA primers. And these two primers are going to be complementary to the opposite strands of D N A and are going to be oriented towards each other and serve as the starting 0.4. The amplification and so down below. What you can see is part B. Here are the DNA primers, which are going to be small little DNA molecules that are oriented opposite from one another. So notice that one goes from five prime 23 prime left to right and the other goes five prime 23 prime 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 thermo stable D n a preliminary race, and the thermo stable 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 third component, see, you can see that we need a thermo stable DNA preliminaries, which is really, uh, commonly going to be a d n a polymerase called tak preliminaries, which we'll get to talk more about Tak preliminaries as we move forward in our course Now, the fourth and final component here that's going to be needed. Uh, in the PCR Uh, mixture is D here, which is going to be all four Deoxyribonucleic tides are all for 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 four of those DNA nucleotides which you can see over here, which include, uh, 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 going to start off with your template d n A. 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 D N. A. The second cycle will amplify the D N A even more, and the third cycle will amplify the D N. A. Even more and continuous cycles will amplify the D N a 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 two raised to the power of n, where N is going to 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 d N A. In the first cycle of PCR, all we need to do is take to raise to the power of one, since one is representing a cycle number one, and so to raise to the one is equal to two. And so notice that after the first cycle there are two new copies of DNA Now, after the second cycle, Uh, if we wanted to calculate how many new copies of template DNA that would be you just take to raise to the power of to and to raise to the power of two is equal to four. And so you can see that there are four new copies of the D. N A. After cycle number two of PCR, and the same goes for cycle number three. If we want to calculate the number of new copies of Template D N. A. You just take to raise it to the power of three, which is the number of cycles Cycle three, and to raise to the three is equal to eight. And so you can see that there are eight copies of the D N A. After cycle number three. 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 preliminaries chain reaction or PCR. So I'll see you all in our next video