Development of a whole blood SNP identity assay

Whole genome sequencing (WGS) is becoming more common in diagnostic testing within the NHS. A necessary component of the testing pathway is to confirm the patient’s identity before the results of WGS are reported to the referring clinician, with a profile of multiple single nucleotide polymorphism (SNP) genotypes compared to those identified during WGS.

This study aimed to develop a method by which whole blood-input PCR could be used to generate the SNP profile, as this would allow sample mix-ups during the DNA extraction process to be detected. Whole blood-input PCR is commonly used in forensics but most techniques include a modified DNA polymerase to overcome the inhibitory effect of various whole blood components.

The technique developed within this study aimed to enable whole blood-input PCR without a modified polymerase and allow for a validated genotyping kit to be used. The proof-of-principle that DNA obtained through crude lysis of whole blood can be amplified through simple PCR was demonstrated. 100% concordance was achieved in the SNP profile obtained by diluted lysate inputs when compared with a DNA input from the same sample, but only when loci with a low read depth were filtered out. The remaining number of loci for five of the eight lysate inputs was insufficient for clinical use.

It is promising that 100% concordance was achieved but further optimisation of the crude lysis technique for use with the genotyping kit is required in order to improve read depth. This will allow more loci to be included when generating a SNP profile, increasing confidence that any clinically significant variants detected by WGS are being reported for the correct patient.