PCR

PCR (Polymerase Chain Reaction) is a process in which, under the catalysis of DNA polymerase, a DNA strand serves as a template. Specific primers are used as starting points, and the system is supplemented with dNTPs, Mg2+, extension factors, and amplification enhancers. Through steps like denaturation, annealing, and extension, a complementary DNA strand is replicated in vitro, creating a copy of the template DNA strand. This process can rapidly and specifically amplify any desired DNA sequence outside of a living organism. Currently, PCR technology finds widespread applications in various fields related to molecular biology.

The main components of a PCR reaction system include the template, Taq DNA polymerase, primers, dNTPs, and reaction buffer. Among these, dNTP stands for deoxy-ribonucleoside triphosphate, including dATP, dGTP, dTTP, dCTP, dUTP, and so on. 'd' stands for deoxidize or deoxy, 'N' represents nitrogenous bases, which could be A, T, G, C, U, and 'TP' stands for triphosphate. The quality and concentration of dNTPs are closely related to PCR amplification efficiency. In a PCR reaction, the concentrations of the four dNTPs should be equal (equimolar preparation). If the concentration of any one of them differs significantly from the others (either too high or too low), it can lead to mismatches. The concentration of dNTPs should be within the range of 50-200 μmol/L. Too low a concentration can reduce the yield of PCR products, while too high a concentration can affect PCR amplification.

Selecting high-quality dNTPs is crucial for PCR reactions. 1. The purity of dNTPs should be high. 2. The freeze-thaw stability of dNTPs should be good. 3. The inter-batch stability of dNTPs should be strong. 4. There should be no residues from human or bacterial genomes in dNTPs. 5. dNTPs should not contain residual DNases, RNases, or cutting enzymes.

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PCR

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