Edgar Christina D, Blair Anna, Tate Warren P
Department of Biochemistry, School of Biomedical Sciences, University of Otago, Dunedin, New Zealand.
Department of Pathology, Dunedin School of Medicine, University of Otago, Dunedin, New Zealand.
Methods Mol Biol. 2025;2920:247-256. doi: 10.1007/978-1-0716-4498-0_14.
Genetic variations in the indoleamine 2,3-dioxygenase (IDO2) gene that are commonly found in the general population have been assessed for their frequency in myalgic encephalomyelitis/chronic fatigue (ME/CFS) patients compared with healthy controls. They have potential for being genetic variations that lead to susceptibility to developing ME/CFS following exposure to a triggering stressor like a viral infection or other major stress events. The IDO2 gene encodes an enzyme that is involved in the tryptophan-kynurenine pathway (TKP), and is activated if there are excessive amounts of tryptophan to prevent excessive serotonin production. The TKP pathway through production of NADH is involved in regulating the immune system and likely plays an important role in ME/CFS. A simple method was developed to evaluate the 5 commonly occurring mutations in this gene in ME/CFS patients and to determine if one or more were present at higher frequency than in healthy controls. This might indicate a susceptibility factor for developing ME/CFS. In this chapter we describe the techniques used to isolate peripheral blood mononuclear cells (PBMCs), extract the DNA, and then do touchdown PCR and DNA sequencing for the analysis.
与健康对照相比，已评估了普通人群中常见的吲哚胺2,3-双加氧酶（IDO2）基因的遗传变异在肌痛性脑脊髓炎/慢性疲劳（ME/CFS）患者中的频率。这些遗传变异有可能在接触病毒感染或其他重大应激事件等触发应激源后导致易患ME/CFS。IDO2基因编码一种参与色氨酸-犬尿氨酸途径（TKP）的酶，当色氨酸过量时该酶被激活，以防止血清素产生过多。TKP途径通过产生NADH参与调节免疫系统，并且可能在ME/CFS中起重要作用。已开发出一种简单方法来评估ME/CFS患者中该基因的5种常见突变，并确定其中一种或多种突变的出现频率是否高于健康对照。这可能表明是发生ME/CFS的一个易感因素。在本章中，我们描述了用于分离外周血单核细胞（PBMC）、提取DNA，然后进行降落PCR和DNA测序以进行分析的技术。
