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This ensures that no contaminating DNA from previous PCR reactions is present in the lab, which could otherwise generate false positives. COLD-PCR (co-amplification at lower denaturation temperature-PCR) is a modified protocol that enriches variant alleles from a mixture of wild-type and mutation-containing DNA samples.
Figure 4: Methylation-specific PCR is a sensitive method to discriminately amplify and detect a methylated region of interest using methylated-specific primers on bisulfite-converted genomic DNA. Such primers will anneal only to sequences that are methylated, and thus containing 5-methylcytosines that are resistant to conversion by bisulfite ...
A strip of eight PCR tubes, each containing a 100 μL reaction mixture Placing a strip of eight PCR tubes into a thermal cycler. The polymerase chain reaction (PCR) is a method widely used to make millions to billions of copies of a specific DNA sample rapidly, allowing scientists to amplify a very small sample of DNA (or a part of it) sufficiently to enable detailed study.
Inverse polymerase chain reaction (Inverse PCR) is a variant of the polymerase chain reaction that is used to amplify DNA with only one known sequence. One limitation of conventional PCR is that it requires primers complementary to both termini of the target DNA, but this method allows PCR to be carried out even if only one sequence is ...
Duplex sequencing overview: Duplex tagged libraries containing sequencing adapters are amplified and result in two types of products each originates from a single strand of DNA. After sequencing the PCR products, the generated reads divide into tag families based on the genomic position, duplex tags, and the neighboring sequencing adapter.
[4] [16] The ideal range of PCR cycles is 9–15 and it is more ideal to pool multiple PCR reactions to get enough DNA for sequencing, than to increase the number of cycles for one PCR reaction. [4] [16] The PCR products are purified again using AMPure beads to remove primer dimers and then quantified before being sequenced.