P262 Dna sequencing with Scisco genetics next generation sequencing HLA typing version 5 kit

Abstract

Aim Next-Generation HLA sequencing gives high resolution results, frequently without ambiguous combinations. However, it is still considered laborious and time-consuming. Scisco Genetics Version 5 kit combines indexing and exon-specific amplification in the same step, therefore the tech time is dramatically reduced to 4 h. Reduced processing time is a great advantage especially for high volume labs. Compared with other long-range PCR methods, the Scisco Genetics NGS version 5 method is much simpler and gives more flexibility. Twenty-four samples for 8 loci can be run using Illumina MiSeq Nano kit (24 h run time). Exon coverage was also increased compared with version 4 (Class I Exon 1 to Exon 7 and Class II Exon 1 to Exon 4), therefore the typing ambiguities are reduced greatly. Methods Forty-seven samples with known HLA types were tested using Scisco Genetics NGS version 5 kit on the Illumina MiSeq instrument. Forty-three DNA samples were isolated from blood, one sample from buccal swabs, and three samples from saliva. Two samples with rare HLA types were included in the sample population. The test data was analyzed using GeMS HLA v65.20 analysis software and the typing results were compared with the previous typing data produced by Scisco Genetics NGS version 4 kit and SBT. Results were compared for accuracy and reproducibility of HLA typing, and the ability to detect rare alleles. Presence of preferential amplification, allelic dropout, and null alleles were also evaluated. Results Of the forty-seven samples tested, forty-one samples showed results that are 100% concordance with the previous data obtained by Scisco version 4 kit. The two rare types (C∗12:143 and DRB1∗11:129) were both detected with the version 5 kit. Allele dropout was observed in two samples, and three samples had failure in one or more loci. We did not get any typing from buccal swab DNA, as was the case with previously used methods. Intronic sequence for common null alleles is also included in the analysis software. No null allele population was detected. Conclusions In summary, fast high resolution DNA sequencing results, with few ambiguities, can be produced utilizing the Scisco Genetics Version 5 NGS HLA Typing Kit.