ABSTRACT
The heat-stable DNA polymerases used in the PCR amplification generally work efficiently only with a DNA template and cannot be used to directly amplify RNA molecules. As is the case with the standard PCR amplification process, the product of the RT/PCR process is a double-stranded DNA molecule of defined size that can be directly examined by gel electrophoresis. For many applications, the yields of DNA obtained through PCR amplification are sufficient to circumvent the necessity of cloning in a vector and biological amplification of the desired DNA fragment. Manipulation of the sequence of the primers, the ions in the reaction (particularly Mg), the annealing temperature, and the rate at which reaction conditions change (the ramp times) during the anneal-elongate-denature cycles have all been found to influence efficiency and specificity of the PCR amplification process.
