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Noninvasive fetal genotyping of paternally inherited alleles using targeted massively parallel sequencing in parentage testing cases
Author(s) -
Yang Donggui,
Liang Hao,
Gao Yu,
Lin Shaobin,
He Zhiming,
Gao Jun,
Sun Hongyu,
Li Qing,
Ma Xiaoyan,
Ou Xueling
Publication year - 2017
Publication title -
transfusion
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.045
H-Index - 132
eISSN - 1537-2995
pISSN - 0041-1132
DOI - 10.1111/trf.14077
Subject(s) - massive parallel sequencing , genotyping , allele , biology , genetics , genotype , pyrosequencing , dna sequencing , cell free fetal dna , allele frequency , fetus , computational biology , prenatal diagnosis , dna , gene , pregnancy
BACKGROUND Researchers have sought to develop a noninvasive protocol for paternity analysis that uses fetal cell‐free DNA (cfDNA) in maternal plasma. Massively parallel sequencing (MPS) is expected to overcome this challenge because it enables the analysis of millions of DNA molecules at a single‐base resolution. STUDY DESIGN AND METHODS Seven women were involved in prenatal paternity testing cases. Before conventional invasive procedures, cfDNA was isolated from maternal plasma. Fetal tissues were then collected, as were blood samples from the alleged fathers. A custom array was designed that targeted 1497 regions containing single‐nucleotide polymorphisms. These regions were massively parallel sequenced. RESULTS In these seven cases, the mean nonmaternal allele fractions in maternal plasma ranged from 3.22% to 6.17%. Setting the allele fraction cutoff of 2.5%, 300 to 491 loci were considered informative for paternal origin and no genetic incompatibilities with the alleged fathers were found. These results were concordant with those of conventional short tandem repeat genotyping. Validation results performed using fetal samples showed that sequencing noise was completely filtered out, and 78.35% to 99.19% of the paternal alleles were accurately genotyped. The fetal cfDNA concentrations ranged from 7.12% to 13.81%, and the overall sequencing error rates ranged from 0.40% to 0.93%. CONCLUSION In our study, we evaluate a straightforward method that can be used to identify paternal alleles based on analyses of paternal alleles and sequencing errors in maternal plasma. Our results support the notion that an MPS‐based method could be utilized in noninvasive fetal genotyping and prenatal paternity analyses.