Development Of Class I Hla Gene In-House Hla-Typing Methods (Hla A, Hla B, And Hla C) Using Next Generation Sequencing Illumina Miseq

Human leukocyte antigen (HLA) is a 3.8 Mb protein presenting antigen whose genetic locus is located in chromosome 6p21 area and have association with more than 100 different diseases that are mostly autoimmune diseases. HLA-typing process using Sanger sequencing still creates ambiguity in the determination of alleles, low-throughput, and costly as it requires a large quantity of sample. Next generation sequencing (NGS) is a method that can overcome the drawbacks of Sanger sequencing. MiSeq Illumina is one of the NGSs that are used for HLA-typing. This study was conducted at the Laboratory of Molecular, Universitas YARSI in a period from May to December 2014. MiSeq provides convenience and flexibility in the preparation methods that can be developed according to the needs of the research laboratory. The results showed that there were a total of 546 SNPs that were heterozygous, 888homozygous SNP, 25 insertions and 23 deletions from the overall 11 amplicon samples with an average coverage with 2x25 read length of 2,106,536x. Our protocol generates good result as we combined long PCR amplicon and size selection method to 300-600 bp fragment.