In this video, we're going to talk more about the second step of each PCR cycle, which is annealing. In this step, the DNA primers actually anneal to the heat-denatured single-stranded DNA. The temperature is lowered to about 55 degrees Celsius. If you recall in step number 1, the temperatures were as high as 95 degrees Celsius, so there is a significant drop from 95 degrees Celsius down to about 55 degrees Celsius, which is important for lowering the temperature. This lowering of the temperature allows for complementary base pairing between the DNA primers and the single-stranded DNA.
Taq polymerase remains inactive at these cooler temperatures because the temperatures are too cool for it to synthesize DNA. When we look at our image on the left hand side, at the second step, annealing, you can see that the temperatures are cooled. Cooler temperatures allow the DNA primers to anneal to the DNA. After the first step of PCR, which is denaturation, we know that we've generated single-stranded DNA, this heat-denatured DNA. In the second step, the temperatures are cooled so that the DNA primers can anneal to the DNA as we see here. Notice that they are annealed on different DNA strands, and they are annealed facing toward each other in terms of their 5-prime and 3-prime ends. This one goes from left to right, and this one goes from right to left, facing toward each other.
Now that these DNA primers have been annealed, in the third step, they can be extended. However, it cannot be extended here in the second step because the temperature of 55 degrees Celsius is just too cold for the Taq polymerase to work. Notice the Taq polymerase is saying, "brrrr, I can't work like this," and he's all cold, and he can only work when we increase those temperatures just a little to the right temperatures. This concludes our brief introduction to the second step of PCR, the annealing of the DNA primers. We'll be able to talk about the 3rd and final step of PCR in our next video.