Failure to genotype Human Cytomegalovirus by PCR-RFLP method due to sequence variation within the primer binding site

Polymerase chain reaction-based restriction fragment length polymorphism (PCR-RFLP) is one of the rapid methods for genotyping Human Cytomegalovirus (HCMV). When genotyping clinical samples by a sensitive nested PCR-based RFLP method for the glycoprotein B (gB) gene of HCMV, it was found that some of the clinical specimens did not give an amplification signal. Analysis of the prototype sequences of the different genotypes showed base pair mismatches over the primer binding site. An alternative assay is suggested for genotyping of HCMV.