Why we use DNA primer instead of RNA primers in the pcr?
Why can't the cell use DNA primers instead of RNA primers?
Perhaps in the cells, RNA primer increases the accuracy of the replication. But how?
Hi Mehdi,
I think that many answers to your questions are more philosophic than scientific, considering what we know by now. But, before I start to talk about that, let's take the easier one: "Why we use DNA primers instead of RNA primers in the PCR?".
That's because DNA is far more stable than RNA, chemically speaking. The additional "-OH" termination at the RNA penthouses makes it more reactive to anything. That's why cells which replace RNA with DNA probably had more success in evolutionary history.
This, actually brings us to question two: "Why can't the cell use DNA primers instead of RNA primers?". Based on RNA reactiveness, some theories about first replicant molecules assume the RNA as the first "living being" on earth, at list regarding the capacity of auto-replication. Thus, we could assume that cells just have conserved the first enzymes responsible for replication, which were (are) able to handle RNA.
I know it doesn't answer your question completely, but it brings us to the next step of this reflection, which is about efficiency. It's reasonable that time is a key role for unicellular cells (that should be our first versions from the past), so faster is better, right? What we do know is that the DNA-polymerase is 10-fold faster than RNA-pol. Therefore, a mutant having a DNA-pol will be 10-fold more abundant than the wild one, that just has an RNA-pol. In other words, it's not about the cells can't use DNA as primers, but cells have been selected to be more efficient replicating from RNA to DNA. There's actually a lot about this, why cells have shifted it, how the mechanisms evolved, etc... You can find it elsewhere.
About your last question, since cells shifted from RNA to DNA as the main genetic molecule, it should be possible that they have to handle the inability of DNA-pols start by itself without a free 3'OH terminus. Likewise, with the increasing of genomes sizes, cells with multiples replication starting-points have got a time-efficient advantage over that ones without it. Therefore, multiples "RNA primers" or "Okazaki fragments" can reduce their replication time exponentially. This way, cells can increase their efficiency in terms of time-consuming. It also possible that the current system we have with multiple enzymes for replication (several RNA and DNA-pols versions) is the best one for the kind of organisms we (Metazoa) become, at least the best system we have tried considering our timeline. So what I could tell you is that today RNA primers increase the accuracy of DNA replication because it allows having other core versions of DNA-pol specialized in different tasks, like extend molecules faster or review its errors.
I hope it helps you someway and sorry if it's not what you were looking for.
Cheers!
Hi Mehdi,
the difference is not philosophic, but constitutive due to difference between DNA and RNA (for higher species as human). the difference between both targets are the presence of introns in DNA, whereas there is no exon in RNA. designing primers for qRT-PCR or transcription analysis need that one of the primers will be across a splice junction, just to avoid contaminated DNA amplification. when using DNA as template, you can design your primers everywhere you want, just following some rules to keep PCR efficiency and specificity in higher rates.
fred
I'm sorry Mehdi and Frederic... I never noticed the specificity behind the questions and the underlying assumption about its RT-qPCR applications. Of course, differences between DNA and RNA aren't philosophical and when I mentioned that, I was clearly talking about the evolution of the mechanisms relative to the questions: "why the cell can't use DNA as a primer?" and "RNA primers accuracy in replication process".
Thank you for the contribution
Cheers.
I agree with Wilson and Frederic and also would like to add that as PCR usees Taq polemerase (DNA polemeras) hence we prefer DNA primer for that enzyme to work. Second, the DNA primer is more temperature stable than RNA primer. So, DNA primer is used for efficient annealing in a continuous temperature dependent invitro reactions like PCR.
Primer RNA is RNA that initiates DNA synthesis. Primers are required for DNA synthesis because no known DNA polymerase is able to initiate polynucleotide chain reaction.
We use DNA primer in PCR, because naturally RNA molecule is not stable like DNA. The answer for second question is that cellular replication use RNA primer because on availability of DNA primer in cellular replication.
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