![]() This technique has been designated as Touch-Up (TU) gradient PCR. Therefore, we report here a PCR amplification technique especially for bisulfite-modified DNA, which requires minimum optimization and produces specific products using a temperature range throughout the amplification process. In addition, some methods require double amplification, using nested primers, which increases the chances of nonspecific product generation. Irrespective of the amplification method used, all require prior optimization of the PCR to determine suitable annealing temperatures. Conventional PCR is rarely used, and specialized modifications of PCR, such as touch-down or nested PCRs, are used frequently. ![]() 4 However, the successful amplification of bisulfite-modified DNA is often challenging. These include methylation-specific PCR, 1 combined bisulfite restriction analysis, 2 methylation-sensitive single nucleotide primer extension, 3 and bisulfite sequencing. Some of these techniques require amplification of the bisulfite-modified DNA to study methylation at specific loci within the genome. Bisulfite-modified DNA is the primary requirement for almost all DNA methylation analysis methods available today.
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