PCR and DNA replication are two processes responsible for DNA synthesis. The enzyme responsible for DNA synthesis in PCR is a thermophilic DNA polymerase such as Taq polymerase while the enzyme responsible for DNA replication is DNA polymerase. Moreover, PCR uses DNA primers while DNA replication uses RNA primers synthesized by RNA primase.
Key Areas Covered
1. What is PCR
– Definition, Process, Importance
2. What is DNA Replication
– Definition, Process, Importance
3. What are the Similarities Between PCR and DNA Replication
– Outline of Common Features
4. What is the Difference Between PCR and DNA Replication
– Comparison of Key Differences
DNA Polymerase, DNA Replication, PCR (Polymerase Chain Reaction), Primers, Taq Polymerase
What is PCR
PCR (polymerase chain reaction) is a widely used molecular biological technique to produce thousands to millions of copies of a particular DNA fragment through the exponential amplification. It was developed by Kary Mullis in 1983. The most significant feature of PCR is that it relies on thermal cycling. Therefore, it permits different temperature-dependent reactions to occur, including DNA melting and enzyme-driven DNA polymerization. On the other hand, the two main reagents used in PCR are the DNA primers, which are complementary to the target sequence and a heat-stable DNA polymerase such as Taq polymerase, isolated from the thermophilic bacterium Thermus aquaticus. Meanwhile, the forward and reverse PCR primers flank the region to be polymerized on the DNA fragment.
Furthermore, the three main steps involved in a PCR are:
- Denaturation – Melting DNA duplex into two single strands by heating to 94-95 °C.
- Annealing – The binding of the forward and reverse primers to the complementary sequences on the template. The temperature of this step depends on the melting temperature of the primer combination.
- Primer extension – DNA polymerase enzyme extends each of the primers at their 3’end by adding complementary bases to the growing strand. The optimum temperature of Taq polymerase, 72 °C is used as the temperature in the extension step. The time of the extension depends on the number of base pairs in the template strand.
Generally, the three steps are repeated for 30-40 times during the PCR to obtain an exponential growth of the DNA fragment of interest.
What is DNA Replication
DNA replication is the biological process responsible for the synthesis of two identical copies of DNA from one copy. Moreover, it is the basis of biological inheritance and aids cells to undergo cell division. Additionally, one of the main features of DNA replication is semiconservative replication. Each DNA strand in the DNA duplex serves as a template for the synthesis of a new DNA strand, which is complementary to it. Furthermore, a set of proteins take part in DNA replication. Basically, they are DNA helicase to unwind DNA duplex, DNA polymerase to polymerize DNA, DNA clamp to prevent the DNA polymerase dissociation, single-strand binding proteins to stabilize single-stranded DNA, topoisomerase to relax DNA strands during polymerization, DNA ligase to ligate Okazaki fragments, primase to synthesize RNA primers, and telomerase to lengthen telomeric DNA.
Moreover, DNA replication is a continuous process, and the three steps in DNA replication are:
- Initiation – Starting DNA replication at the origin of replication with the help of origin recognition complex.
- Elongation – Synthesis of DNA in the 5′ to 3′ direction on both leading and lagging strand by DNA polymerase. During elongation, the replication fork is formed. Also, the polymerization on the leading strand is continuous, and the polymerization on the lagging strand occurs through the formation of Okazaki fragments.
- Termination – Blockage of DNA replication by a combination of a termination site sequence in the DNA, and a protein which binds to this sequence to physically stop DNA replication.
Similarities Between PCR and DNA Replication
- PCR and DNA replication are two processes of DNA synthesis.
- Both are polymerizing chain reactions.
- Furthermore, they proceed in the 5′ to 3′ direction in each strand.
- Therefore, the polymerization of the two DNA strands, which are antiparallel occurs in opposite directions.
- Also, DNA polymerizing enzymes carry out both processes.
- The main function of these enzymes to add complementary bases to the growing strand.
- Moreover, both processes use primers, which are short oligonucleotide fragments complementary to DNA.
- Primers are responsible for the initiation of DNA synthesis.
- Both processes use existing DNA as the DNA template for the synthesis of new DNA.
- Therefore, they occur in a semiconservative manner.
- They use deoxyribose nucleotides as substrates.
Difference Between PCR and DNA Replication
PCR (polymerase chain reaction) refers to a method widely used in molecular biology to make many copies of a specific DNA segment while DNA replication refers to the biological process of producing two identical replicas of DNA from one original DNA molecule. Thus, this is the main difference between PCR and DNA replication.
PCR is an in vitro process, which occur inside a test tube while DNA replication is an in vivo process, which occur inside living cells.
The main goal of PCR is to produce 230 to 240 copies of a single DNA fragment while the main goal of DNA replication is to copy the whole genome at once.
The target of PCR is shorter while the target of DNA replication is longer.
Continuity of the Process
PCR is a discontinuous process which proceeds through 30-40 cycles while DNA replication is a continuous process.
Opening up the Two Strands of DNA Helix
In PCR, DNA duplex is melted by using heat, which is >90 °C while in DNA replication, DNA duplex is opened up by the enzyme ATP-dependent helicase.
Another difference between PCR and DNA replication is that PCR uses DNA primers while DNA replication uses RNA primers synthesized by primase enzyme.
Moreover, PCR uses thermophilic DNA polymerase such as Taq DNA polymerase while DNA replication uses DNA polymerase.
Features of Polymerases
Taq polymerase in PCR is not feature-rich, and also, it has no proofreading ability while DNA polymerase in DNA replication is contained high fidelity, speed, proofreading and repair abilities.
Replication fork does not occur in PCR while replication fork occurs in DNA replication.
5′ to 3′ Exonuclease Activity
Taq polymerase in PCR does not contain the 5′ to 3′ exonuclease activity while DNA polymerase in DNA replication has the 5′ to 3′ exonuclease activity to degrade RNA primers.
Taq polymerase in PCR operates at high temperatures such as 72 °C while DNA polymerase in DNA replication operates at physiological temperature, which is 37 °C.
PCR serves as a simple approach for in vitro DNA synthesis while DNA replication is a complex process, which depends upon a well defined but complex set of enzymes and co-factors.
The speed of PCR is 1-4 kb/min while the speed of DNA replication is 1 kb/s.
The error rate of Taq polymerase in PCR is 1 in 9000 bases while the error rate of DNA polymerase in DNA replication is 1 in 100,000 bases.
Basically, PCR is the process of in vitro DNA synthesis. Also, it is a simple approach which uses high temperatures and thermophilic Taq polymerase as the enzyme. Additionally, it uses DNA primers. However, the main goal of PCR is to produce a large number of copies of a particular DNA fragment. On the other hand, DNA replication is the in vivo process of DNA synthesis responsible for the synthesis of the whole genome, preparing the cell to divide. However, it requires a number of physiological agents along with the enzyme DNA polymerase. Moreover, this enzyme operates at body temperature. Additionally, DNA replication is a highly accurate process. Therefore, the main difference between PCR and DNA replication is the different features of the process.
1. “Polymerase Chain Reaction (PCR).” Khan Academy, Khan Academy, Available Here.
2. “What Is DNA Replication?” Yourgenome, The Public Engagement Team at the Wellcome Genome Campus, 25 Jan. 2016, Available Here.