Kanthi Kariyawasam

Killer Cell Immunoglobulin-Like Receptors (KIR) Typing By DNA Sequencing

DNA sequencing is a powerful technique for identifying allelic variation within the natural killer cell immunoglobulin-like receptor genes. Because of the relatively large size of the KIR genes, each locus is amplified in two or more overlapping segments. Sanger sequencing of each gene from a preparation containing one or two alleles yields a sequence that is used to identify the alleles by comparison with a reference database.

Extensive haplotype diversity in African American mothers and their cord blood units.

HLA-A, -B, -C, -DRB1, -DQB1 assignments were obtained for 374 pairs of African American mothers and their umbilical cord blood units (CBU) by DNA sequencing. An algorithm developed by the National Marrow Donor Program was used to assign 1122 haplotypes by segregation. Seventy percent of the haplotypes carried assignments at all five loci. In the remainder, alleles at various loci, most often DQB1 in 48% of the haplotypes with a missing assignment, could not be assigned due to sharing of both alleles by mother and CBU. There were 652 haplotypes carrying a unique combination of alleles at the five loci; the majority (74%) were singletons. Novel B∼C and DRB1~DQB1 associations were observed. The results show the genetic diversity in this population and provide validation for a publically available tool for pedigree analysis. Our observations underscore the need for procurement of increased numbers of units in the national cord blood inventory in order to identify matching donors for all patients requiring hematopoietic stem cell transplantation.

Combining one‐step Sanger sequencing with phasing probe hybridization for HLA class I typing yields rapid, G‐group resolution predicting 99% of unique full length protein sequences

Sanger‐based DNA sequencing of exons 2+3 of HLA class I alleles from a heterozygote frequently results in two or more alternative genotypes. This study was undertaken to reduce the time and effort required to produce a single high resolution HLA genotype.

Full gene HLA class I sequences of 79 novel and 519 mostly uncommon alleles from a large United States registry population

HLA class I assignments were obtained at single genotype, G‐level resolution from 98 855 volunteers for an unrelated donor registry in the United States. In spite of the diverse ancestry of the volunteers, over 99% of the assignments at each locus are common. Within this population, 52 novel alleles differing in exons 2 and 3 are identified and characterized. Previously reported alleles with incomplete sequences in the IPD‐IMGT/HLA database (n = 519) were selected for full gene sequencing and, from this sampling, another 27 novel alleles are described.