Haplotype-Based noninvasive prenatal diagnosis for duchenne muscular dystrophy: A pilot study in South China

AbstractObjectiveWe aimed to investigate the validity of noninvasive prenatal diagnosis (NIPD) based on direct haplotype phasing without the proband and its feasibility for clinical application in the case of Duchenne Muscular Dystrophy (DMD).MethodsThirteen singleton-pregnancy families affected by DMD were recruited. Firstly, we resolved maternal haplotypes for each family by performing targeted linked-read sequencing of their high molecular weight DNA, respectively. Then, we identified SNPs of the DMD gene in all carrier mothers and inferred the DMD gene mutation status of all fetuses. Finally, the fetal genotypes were further validated by using chorionic villus sampling.ResultsThe method of directly resolving maternal haplotype through targeted linked-read sequencing was smoothly performed in all participated families. The predicted mutational status of 13 fetuses was correct, which had been confirmed by invasive prenatal diagnosis.ConclusionDirect haplotyping of NIPD based on linked-read sequencing for DMD is accurate.

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Haplotype-Based Noninvasive Prenatal Diagnosis for Duchenne Muscular Dystrophy: A pilot study in South China

10.1101/551200 ◽

2019 ◽

Author(s):

Min Chen ◽

Chao Chen ◽

Yingting Li ◽

Yuan Yuan ◽

Zhengfei Lai ◽

...

Keyword(s):

Prenatal Diagnosis ◽

Duchenne Muscular Dystrophy ◽

Muscular Dystrophy ◽

Maternal Plasma ◽

Targeted Sequencing ◽

Plasma Dna ◽

Noninvasive Prenatal Diagnosis ◽

Integration Analysis ◽

Dependent Probe ◽

Singleton Pregnancies

AbstractObjectiveTo explore the accuracy and feasibility of noninvasive prenatal diagnosis (NIPD) for Duchenne Muscular Dystrophy (DMD) based on the haplotype approach.MethodsWe recruited singleton pregnancies at-risk of DMD at 12-25 weeks of gestation from 17 families who all had a proband children affected by DMD. We have identified the pathogenic mutations in probands and their mothers by multiplex ligation-dependent probe amplification (MLPA). To construct parental haplotypes, we performed captured sequencing on genomic DNA from parents and probands. The integration analysis of parental haplotypes and targeted sequencing results of maternal plasma DNA were used to infer the fetal haplotype and genotypes in DMD gene. Fetal DMD genotypes were further confirmed by invasive prenatal diagnosis.ResultsWe have successfully performed the haplotype-based NIPD in all recruited families. Ten fetuses were identified as normal, including four female and six male fetuses. Four female fetuses were carriers and the other three male fetuses were affected by DMD with exons 49-52 deletion, exons 8-37 deletion and c.628G > T mutation, respectively. The results of NIPD were consistent with those of invasive diagnosis.ConclusionHaplotype-based NIPD for DMD by targeted sequencing is promising and has potential for clinical application.

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Haplotype-Based Noninvasive Prenatal Diagnosis of 21 Families With Duchenne Muscular Dystrophy: Real-World Clinical Data in China

Frontiers in Genetics ◽

10.3389/fgene.2021.791856 ◽

2021 ◽

Vol 12 ◽

Author(s):

Lingrong Kong ◽

Shaojun Li ◽

Zhenhua Zhao ◽

Jun Feng ◽

Guangquan Chen ◽

...

Keyword(s):

Prenatal Diagnosis ◽

Duchenne Muscular Dystrophy ◽

Muscular Dystrophy ◽

Family Members ◽

Chorionic Villus ◽

Genetic Diagnosis ◽

Blood Collection ◽

Chorionic Villus Sampling ◽

Clinical Environment ◽

Noninvasive Prenatal Diagnosis

Noninvasive prenatal diagnosis (NIPD) of single-gene disorders has recently become the focus of clinical laboratories. However, reports on the clinical application of NIPD of Duchenne muscular dystrophy (DMD) are limited. This study aimed to evaluate the detection performance of haplotype-based NIPD of DMD in a real clinical environment. Twenty-one DMD families at 7–12 weeks of gestation were prospectively recruited. DNA libraries of cell-free DNA from the pregnant and genomic DNA from family members were captured using a custom assay for the enrichment of DMD gene exons and spanning single-nucleotide polymorphisms, followed by next-generation sequencing. Parental haplotype phasing was based on family linkage analysis, and fetal genotyping was inferred using the Bayes factor through target maternal plasma sequencing. Finally, the entire experimental process was promoted in the local clinical laboratory. We recruited 13 complete families, 6 families without paternal samples, and 2 families without probands in which daughter samples were collected. Two different maternal haplotypes were constructed based on family members in all 21 pedigrees at as early as 7 gestational weeks. Among the included families, the fetal genotypes of 20 families were identified at the first blood collection, and a second blood collection was performed for another family due to low fetal concentration. The NIPD result of each family was reported within 1 week. The fetal fraction in maternal cfDNA ranged from 1.87 to 11.68%. In addition, recombination events were assessed in two fetuses. All NIPD results were concordant with the findings of invasive prenatal diagnosis (chorionic villus sampling or amniocentesis). Exon capture and haplotype-based NIPD of DMD are regularly used for DMD genetic diagnosis, carrier screening, and noninvasive prenatal diagnosis in the clinic. Our method, haplotype-based early screening for DMD fetal genotyping via cfDNA sequencing, has high feasibility and accuracy, a short turnaround time, and is inexpensive in a real clinical environment.

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Noninvasive prenatal diagnosis of duchenne muscular dystrophy in five Chinese families based on relative mutation dosage approach

BMC Medical Genomics ◽

10.1186/s12920-021-01128-1 ◽

2021 ◽

Vol 14 (1) ◽

Author(s):

Ganye Zhao ◽

Xiaofeng Wang ◽

Lina Liu ◽

Peng Dai ◽

Xiangdong Kong

Keyword(s):

Prenatal Diagnosis ◽

Duchenne Muscular Dystrophy ◽

Muscular Dystrophy ◽

Single Molecule ◽

De Novo ◽

Dna Barcode ◽

De Novo Mutations ◽

Molecular Alteration ◽

Noninvasive Prenatal Diagnosis ◽

Exon Deletion

Abstract Background Relative haplotype dosage (RHDO) approach has been applied in noninvasive prenatal diagnosis (NIPD) of Duchenne muscular dystrophy (DMD). However, the RHDO procedure is relatively complicated and the parental haplotypes need to be constructed. Furthermore, it is not suitable for the diagnosis of de novo mutations or mosaicism in germ cells. Here, we investigated NIPD of DMD using a relative mutation dosage (RMD)-based approach—cell-free DNA Barcode-Enabled Single-Molecule Test (cfBEST), which has not previously been applied in the diagnosis of exon deletion. Methods Five DMD families caused by DMD gene point mutations or exon deletion were recruited for this study. After the breakpoints of exon deletion were precisely mapped with multiple PCR, the genotypes of the fetuses from the five DMD families were inferred using cfBEST, and were further validated by invasive prenatal diagnosis. Results The cfBEST results of the five families indicated that one fetus was female and did not carry the familial molecular alteration, three fetuses were carriers and one was male without the familial mutation. The invasive prenatal diagnosis results were consistent with those of the cfBEST procedure. Conclusion This is the first report of NIPD of DMD using the RMD-based approach. We extended the application of cfBEST from point mutation to exon deletion mutation. The results showed that cfBEST would be suitable for NIPD of DMD caused by different kinds of mutation types.

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