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SEPARATION AND SNP-TYPING OF GENOMIC DNA BY HAPLOTYPE-SPECIFIC EXTRACTION AND TAQMAN.
Eleni E. Magira, MD, Johannes Dapprich, PhD, Colleen Murphy, MT, Dylan Membrino, MT, Cynthia Turino, MT, Nancy Murphy, MT, Robert Baldassano, MD and Dimitri S. Monos, PhD. Philadelphia PA, USA, CHOP/UPENN, 19104, Immunogenetics; Lawrenceville PA, USA, Generation Biotech; West Chester USA, GenoVision and Philadelphia PA, USA, CHOP, 19104, Center for Inflammatory Bowel Diseases.

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Haplotype-Specific Extraction, HSE, establishes haplotypes from individual samples by physically separating a diploid sample into its haploid components: Magnetic beads are selectively attached to polymorphic sites and used to isolate the targeted fragments of genomic DNA from a heterozygous mixture. We have used Haplotype-Specific Extraction / HaploPrepô and TaqManôto further characterize MHC-haplotypes based on seven SNP-markers located in a 100kb region of HLA-B and -C. Four families, have been typed and their haplotypes were derived based on diploid HLA-B, -C as well as seven polymorphic SNP sites. Independently, the diploid samples were haplo-separated based on known heterozygous sites at HLA-B or -C, as confirmed by InnoLiPA typing. We established the SNP-haplotypes directly by TaqMan-typing the haploid samples. The results confirmed the family-based haplotype structure. This demonstrates that HSE / HaploPrepô and TaqManôcan extend the analysis of MHC haplotypes to other HLA-linked genes and polymorphic markers such as micro-satellites and SNPs. This approach eventually may be useful to study disease association questions.