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Characterization of killer immunoglobulin-like receptor genetics and comprehensive genotyping by pyrosequencing in rhesus macaques

Anna J Moreland1, Lisbeth A Guethlein2, R Keith Reeves3, Karl W Broman4, R Paul Johnson3, Peter Parham2, David H O'Connor15 and Benjamin N Bimber5*

Author Affiliations

1 Department of Pathology and Laboratory Medicine, University of Wisconsin-Madison, Madison, Wisconsin 53706, USA

2 Departments of Structural Biology and Microbiology & Immunology, Stanford University School of Medicine, Stanford, CA 94305, USA

3 Division of Immunology, New England Primate Research Center, Harvard Medical School, Southborough, MA 01772, USA

4 Department of Biostatistics and Medical Informatics, University of Wisconsin-Madison, Madison, Wisconsin 53706, USA

5 Wisconsin National Primate Research Center, University of Wisconsin-Madison, Madison, Wisconsin 53706, USA

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BMC Genomics 2011, 12:295  doi:10.1186/1471-2164-12-295

Published: 7 June 2011

Additional files

Additional file 1:

Figure S1. Alignment of all published rhesus macaque lineage II KIR sequences. An alignment was generated from all previously published rhesus macaque KIRs using predicted amino acid sequences. Sequences identified in this publication are highlighted. KIRs are grouped by gene, and a consensus sequence is included for each gene. An asterisk after the accession number indicates this sequence was published multiple times and only one of the accession numbers is given. Table S1. Genbank accession numbers for novel full-length rhesus macaque KIR sequences. Sequences have been assigned official names through the Immuno Polymorphism Database. Table S2. PCR primers used for cDNA-PCR pyrosequencing. Table S3. Comparison of pyrosequencing and cloning results. For each animal, detected KIR alleles are shown, along with their relative frequency detected by pyrosequencing, expressed as a percent of total 454 reads. The column on the right indicates the result of conventional cloning. A plus sign indicates that the detected resolution matches the corresponding pyrosequencing result. If cloning resulted in a different resolution, the corresponding allele name is shown. Table S4. Reproducibility of allele frequency estimates. Pyrosequencing data from four animals are shown. Two independent NK cell isolations were performed per animal, and two independent PCRs were performed per cell preparation, with the exception of 225-07, for which only one cell pellet was available. The resulting PCR amplicons were pyrosequenced. The total number of reads is shown for each reaction. KIRs detected are shown, expressed as a percent of total reads. Table S5. GenBank accession numbers for novel partial length rhesus macaque KIR sequences identified by pyrosequencing. Sequences have been assigned sequential, unofficial names. For each sequence, the KIR allele or gene to which it bears greatest similarity is indicated. The Total Ids column indicates the number of distinct animals in which that sequence was observed. Abbreviations: u: unique; gc: gene conversion; r: recombination; sv: splice variant.

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