**2.2. Polymerase chain reaction related ambiguities**

Polymerase chain reaction (PCR) is an essential part of most NGS workflows; in many cases it is a step of the library preparation process (it can be used for targeting and/or amplification) and in all major sequencing platforms PCR is part of the actual sequencing step (emulsion PCR in Ion Torrent and Roche 454 and bridge PCR in Illumina). Considering the major role that PCR plays in NGS, it is important to be aware of possible errors and artifacts that can originate from PCR, as these can greatly affect the outcome of HLA genotyping. PCR-related ambiguities are usually caused by two issues:

