qPCR is the preferred choice for quantifying NGS libraries due to its ability to precisely measure amplifiable molecules. It offers higher sensitivity, requires less sample input, and can be scaled up for high-throughput applications.
Spectrophotometry measures the absorption of light by a substance as a function of wavelength. In nucleic acid analysis, spectrophotometry is used to determine the concentration and purity of the extracted DNA or RNA sample. The two key measurements obtained from spectrophotometric analysis are the absorbance at 260 nm (A260) and the ratio of absorbance at 260 nm to 280 nm (A260/A280). The absorbance at 260 nm is directly proportional to the concentration of nucleic acids in the sample.
Gel electrophoresis is a technique used to separate and visualize nucleic acids based on their size and charge. In this method, the nucleic acid samples are loaded onto an agarose or polyacrylamide gel and subjected to an electric field. Smaller DNA or RNA fragments travel faster through the gel matrix than larger fragments, resulting in separation according to size. Gel electrophoresis provides an estimation of the quantity of nucleic acids present in the sample based on the intensity of the bands observed on the gel.