Identification of novel alleles and structural haplotypes of major histocompatibility complex class I and DRB genes in domestic cat (Felis catus) by a newly developed NGS-based genotyping method. Felis catus

Peripheral white blood cells (PWBCs) were obtained from 20 related domestic cats from two families and five unrelated cats (A6, A116, A165, A176 and A214). Each family was composed of ten cats representing three generations that were considered suitable for investigating the FLA allele and haplotype segregation. We first used the five unrelated cats to elucidate the alleles and specificity of expressed FLA-I loci in PWBCs and then used the 20 related cats for the NGS-based genotyping of expressible FLA-I and FLA-DRB loci. We have identified and presented novel alleles for the FLA-I and FLA-DRB genes and provided some further insights into the development of the NGS-based genotyping method at the RNA expression level, phylogenetic relationships and haplotype structures of FLA-I and FLA-DRB alleles and haplotypes in 20 cats from two families. This FLA polymorphism information could be useful as a standard internal control to establish the feline MHC genetic background for the benefit of biomedical research into disease associations such as infectious disease and autoimmune diseases in veterinary medical field.

外周血白细胞（Peripheral white blood cells, PWBCs）采集自两个家系的20只近缘家猫，以及5只无关家猫（编号A6、A116、A165、A176及A214）。每个家系包含10只涵盖三代的家猫，该家系结构适用于研究猫白细胞抗原（Feline Leukocyte Antigen, FLA）等位基因与单体型的分离情况。本研究首先利用5只无关家猫，阐明了外周血白细胞中表达型FLA-I基因座的等位基因及其特异性；随后借助20只近缘家猫，针对可表达的FLA-I与FLA-DRB基因座开展基于下一代测序（Next-Generation Sequencing, NGS）的基因分型工作。本研究已鉴定并报道了FLA-I与FLA-DRB基因的新型等位基因，并从RNA表达水平的NGS基因分型方法开发、两个家系20只家猫的FLA-I及FLA-DRB等位基因与单体型的系统发育关系、单体型结构等维度，提供了全新的研究视角。本研究获得的FLA多态性信息可作为标准内参，用于构建猫科主要组织相容性复合体（Major Histocompatibility Complex, MHC）的遗传背景，从而助力兽医领域中感染性疾病、自身免疫性疾病等疾病关联的生物医学研究。