What are the common challenges in sample preparation for NGS?

One common challenge is that specimens are often extracted from a small number of cells (such as a single cell) and require amplification through PCR due to the limited amount of material available. This amplification process can introduce bias to the sample. Also, PCR duplication occurs when multiple copies of the same DNA fragment are generated, potentially resulting in uneven sequencing coverage. Contamination in samples is another common issue, especially when multiple libraries are prepared simultaneously. The main source of contamination typically stems from pre-amplification. Another issue is during sample preparation, inefficient library construction can occur. This is exhibited by a low proportion of fragments with the right adaptors. As a result, less sequencing data is obtained, and there is a higher occurrence of chimeric fragments. Lastly, the high expenses associated with library preparation primarily arise from the investment in laboratory equipment, and the costs of reagents.