Sources of PCR inhibitors
Mechanism of action of PCR inhibitors
Removal of PCR inhibitors1. The most fundamental way to overcome PCR inhibition is to remove the inhibitor from the sampleA. Sample processing For different types of samples, appropriate pretreatment methods such as digestion, dissolution, washing, etc. are used to remove inhibitors. (1) Plant samples: The CTAB method, SDS method and other extraction methods are used to remove inhibitors such as polyphenols and polysaccharides. (2) Animal samples: Inhibitors such as proteins and lipids can be removed by methods such as proteinase K digestion and phenol-chloroform extraction. (3) Microbiological samples: Inhibitors such as humic acid can be removed by using reagents such as sodium chloride and SDS, combined with steps such as centrifugation and washing. B. Sample Purification Use purification methods such as filtration columns and magnetic beads to remove inhibitors. Second, the simplest way to overcome PCR inhibition is to dilute the sample, thereby reducing the concentration of PCR inhibitors. Third, the most worry-free method is to use a modified DNA polymerase or reaction system that is resistant to PCR inhibitors. For example, by using enzymes with strong tolerance to inhibitors or adding stabilizers such as bovine serum albumin (BSA) to the reaction system, the success of subsequent analysis can be guaranteed. However, the cost of improved enzymes is often high, and different improved enzymes and different optimized reaction systems may be required for different PCR inhibitors. ***To ensure the experimental results, we should avoid introducing new PCR inhibitors when removing PCR inhibitors. For example, in the selection of reagents, we should choose high-quality, inhibitor-free enzyme preparations, dNTPs, primers, etc.; during the experiment, we should strictly follow the experimental operation specifications to ensure that no inhibitors are introduced during the experiment, and ensure that the experimental equipment is fully washed to avoid cross contamination. SummarizePCR inhibitors are the "invisible killers" that affect the success of PCR experiments. Understanding their sources, mechanisms of action, and removal methods is of great significance for us to conduct PCR experiments. By taking a series of measures, such as optimizing sample processing, selecting high-quality reagents, and standardizing experimental operations, we hope to minimize the impact of inhibitors and ensure the smooth progress of the experiment. Related Products
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