An SNP panel for the analysis of paternally inherited alleles in maternal plasma using ion Torrent PGM
An SNP panel for the analysis of paternally inherited alleles in maternal plasma using ion Torrent PGM
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使用 ion Torrent PGM 分析母体血浆中父系遗传等位基因的 SNP panel
DOI：
10.1007/s00414-017-1594-6
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发表时间：
2018-03-01
影响因子：
2.1
通讯作者：
Ou，Xueling
中科院分区：
文献类型：
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作者：
Yang，Donggui;Liang，Hao;Ou，Xueling
研究人员试图开发一种利用母体血浆中的无细胞DNA （cfDNA）进行亲子鉴定的有效方案。大规模平行测序（MPS）技术用于SNP检测因其高通量和单碱基精度的分辨率而具有吸引力。在这项研究中，我们为cfDNA测序设计了一个定制的SNP面板，其中包括720个短扩增子（< 140 bp），靶向常染色体和Y染色体上的SNP。使用Ion Torrent PGM对系统性能进行了评估，结果表明，除了78个表现不佳的snp存在不一致的等位基因平衡、较低的覆盖读数或高背景信号外，大多数被纳入的位点的覆盖都是平衡的。然后，使用定制面板对20例妊娠早期和中期（9至21周）孕妇的血浆进行cfDNA基因分型。通过建立2.0%的等位基因分数截止点，53至128个常染色体SNP位点被认为是父系起源的信息。胎儿样本验证结果显示，真实父本等位基因鉴定准确率为49.43% ~ 100%，其中3例出现错误等位基因。胎儿cfDNA浓度范围为4.28% ~ 10.70%。我们的研究结果表明，这种基于扩增子的测序策略可用于分析母体血浆中的父系遗传等位基因。然而，基于MPS技术的cfDNA测序结果的更详细和准确的解释需要进一步的研究和优化。
Researchers have sought to develop an effective protocol for paternity analysis using cell-free DNA (cfDNA) in maternal plasma. The use of massively parallel sequencing (MPS) technology for SNP testing is attractive because of its high-throughput capacity and resolution to single-base precision. In this study, we designed a customized SNP panel for cfDNA sequencing that includes 720 short amplicons (< 140 bp) targeting SNPs on the autosome and Y chromosome. The systemic performance was evaluated using the Ion Torrent PGM, indicating balanced coverage among most of the included loci, except for 78 poorly performing SNPs that were observed to have an inconsistent allele balance, lower coverage reads or high background signals. Then, the custom panel was used to perform cfDNA genotyping in maternal plasma from 20 pregnancies in the first and second trimesters (9 to 21 weeks). By establishing an allele fraction cutoff of 2.0%, 53 to 128 autosomal SNP loci were considered informative for paternal origin. Validation results in foetal samples showed that 49.43% to 100% of the real paternal alleles were accurately identified, with incorrect alleles encountered in 3 cases. The concentration of foetal cfDNA ranged from 4.28% to 10.70%. Our results show that this amplicon-based sequencing strategy could be utilized in analysing paternally inherited alleles in maternal plasma. However, further studies and optimization are required for a more detailed and accurate interpretation of the cfDNA sequencing results based on MPS technology.