■地中海贫血是单基因的,常染色体隐性遗传,由珠蛋白基因突变或缺失引起的红细胞遗传性疾病。大约6-10%的印度尼西亚人口携带β-珠蛋白基因突变;然而,很少进行婚前筛查,产前筛查是可选的。我们探索了使用无细胞胎儿DNA(cffDNA)作为一种潜在的非侵入性方法来检测孕妇产前胎儿β-珠蛋白基因突变。
■怀孕母亲(n=10),他们是已知的地中海贫血携带者,并且有患有重型地中海贫血的婴儿的历史,他们的携带者丈夫(n=4)在提供同意后被招募。抽取EDTA血液,和母体DNA,包括cffDNA,并分离父系DNA。使用ApoB和D1S80基因座的可变数量串联重复测试进行母体污染测试。通过焦磷酸测序进行等位基因定量。来自重型地中海贫血患者(n=16)的生物存档DNA的已知突变与对照一起运行。
■总共,10个cffDNA中有7个胜利通过母体沾染试验。等位基因定量结果显示,6例胎儿为IVS1nt5的预测携带者,1例预测正常,与重型地中海贫血患者生物存档DNA的等位基因定量一致。cd26突变A等位基因的最小阈值百分比为32%,IVS1nt1突变T等位基因为23%,IVS1nt5突变C等位基因为39%。
■从母亲的血液中获取cffDNA被证明是一种使用焦磷酸测序等位基因定量检测β-珠蛋白基因突变的非侵入性手段。这种非侵入性方法对于设施有限的环境中的产前诊断非常感兴趣。因为它最大限度地降低了堕胎的风险。需要进一步研究β-珠蛋白基因的其他突变。
UNASSIGNED: Thalassemia is a monogenic, autosomal recessive, inherited disorder of the red blood cells caused by mutations or deletions in the globin gene. Approximately 6-10% of the Indonesian population carries the β-globin gene mutation; however, premarital screening is rarely conducted, and antenatal screening is optional. We explored the use of cell-free fetal DNA (cffDNA) as a potential non-invasive method of detecting the fetal β-globin gene mutation prenatally in pregnant women.
UNASSIGNED: Pregnant mothers (n = 10), who were known carriers of thalassemia and who had a history of having borne a baby with thalassemia major, and their carrier husbands (n = 4) were recruited after providing consent. EDTA blood was drawn, and maternal DNA, including cffDNA, and paternal DNA were isolated. Maternal contamination tests were conducted using the variable number tandem repeat test for ApoB and D1S80 loci. Allele quantification was performed by
pyrosequencing. Known mutations from the bio-archived DNA of patients with thalassemia major (n = 16) were run alongside as a control.
UNASSIGNED: In total, 7 out of 10 cffDNA successfully passed the maternal contamination test. The results of the allele quantification showed that six fetuses were predictive carriers of IVS1nt5 and one was predictive normal, in line with the allele quantification for the bio-archived DNA from patients with thalassemia major. The minimum threshold percentage for mutant A allele at cd26 was 32%, mutant T allele at IVS1nt1 was 23%, and mutant C allele at IVS1nt5 was 39%.
UNASSIGNED: Taking cffDNA from the mother\'s blood proved useful as a non-invasive means of detecting the β-globin gene mutation using
pyrosequencing allele quantification. This non-invasive method is of great interest for prenatal diagnosis in settings with limited facilities, as it minimizes the risk of abortion. Further
study of other mutations of the β-globin gene is needed.