Department of Transfusion Medicine, NIH Clinical Center, National Institutes of Health, Bethesda, Maryland, USA.
Department of Pathology and Laboratory Services, ECU Health Medical Center, Greenville, North Carolina, USA.
Transfusion. 2024 Jul;64(7):1296-1305. doi: 10.1111/trf.17869. Epub 2024 May 30.
CD59 deficiency due to rare germline variants in the CD59 gene causes disabilities, ischemic strokes, neuropathy, and hemolysis. CD59 deficiency due to common somatic variants in the PIG-A gene in hematopoietic stem cells causes paroxysmal nocturnal hemoglobinuria. The ISBT database lists one nonsense and three missense germline variants that are associated with the CD59-null phenotype. To analyze the genetic diversity of the CD59 gene, we determined long-range CD59 haplotypes among individuals from different ethnicities.
We determined a 22.7 kb genomic fragment of the CD59 gene in 113 individuals using next-generation sequencing (NGS), which covered the whole NM_203330.2 mRNA transcript of 7796 base pairs. Samples came from an FDA reference repository and our Ethiopia study cohorts. The raw genotype data were computationally phased into individual haplotype sequences.
Nucleotide sequencing of the CD59 gene of 226 chromosomes identified 216 positions with single nucleotide variants. Only three haplotypes were observed in homozygous form, which allowed us to assign them unambiguously as experimentally verified CD59 haplotypes. They were also the most frequent haplotypes among both cohorts. An additional 140 haplotypes were imputed computationally.
We provided a large set of haplotypes and proposed three verified long-range CD59 reference sequences, based on a population approach, using a generalizable rationale for our choice. Correct long-range haplotypes are useful as template sequences for allele calling in high-throughput NGS and precision medicine approaches, thus enhancing the reliability of clinical diagnostics. Long-range haplotypes can also be used to evaluate the influence of genetic variation on the risk of transfusion reactions or diseases.
由于 CD59 基因的罕见种系变异导致的 CD59 缺乏会引起残疾、缺血性中风、周围神经病和溶血性贫血。由于造血干细胞中 PIG-A 基因的常见体细胞变异导致的 CD59 缺乏会引起阵发性夜间血红蛋白尿。ISBT 数据库列出了一个无义突变和三个错义突变种系变异与 CD59 缺失表型相关。为了分析 CD59 基因的遗传多样性，我们在来自不同种族的个体中确定了长程 CD59 单倍型。
我们使用下一代测序（NGS）在 113 个人中确定了 CD59 基因的 22.7kb 基因组片段，该片段覆盖了 7796 个碱基对的整个 NM_203330.2 mRNA 转录本。样本来自 FDA 参考库和我们的埃塞俄比亚研究队列。原始基因型数据被计算性地相形成个体单倍型序列。
对 226 条染色体的 CD59 基因进行测序，确定了 216 个具有单核苷酸变异的位置。仅在纯合形式下观察到三种单倍型，这使我们能够将它们明确地分配为经过实验验证的 CD59 单倍型。它们也是两个队列中最常见的单倍型。通过计算推断出另外 140 种单倍型。
我们基于人群方法提供了一大组单倍型，并提出了三个经过验证的长程 CD59 参考序列，选择的依据是一个可推广的合理理由。正确的长程单倍型可用作高通量 NGS 和精准医疗方法中等位基因调用的模板序列，从而提高临床诊断的可靠性。长程单倍型还可用于评估遗传变异对输血反应或疾病风险的影响。
