A capillary electrophoresis-based multiplex PCR assay for expanded carrier screening in the eastern Han Chinese population

Hu, Ping; Tan, Jianxin; Yu, Feng; Shao, Binbin; Zhang, Fang; Zhang, Jingjing; Lin, Ying‐Chun; Tao, Tao; Jiang, Lili; Jiang, Zhengwen; Xu, Zhengfeng

doi:10.1038/s41525-021-00280-y

Cited by 4 publications

(7 citation statements)

References 55 publications

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“…The price of SNaPshot/HLPA was dramatically lower than that for NGS ($85 per case vs. $170 per case), so it might be widely used in economically underdeveloped regions. Furthermore, turnaround time for SNaPshot/HLPA was 3 days, which is obviously shorter than that for NGS (2 weeks) (Hu et al., 2022).…”

Section: Discussionmentioning

confidence: 99%

“…Genomic DNA samples from peripheral blood leukocytes were extracted by Automated Nucleic Acid Extractor (RBC Bioscience, New Taipei City, Taiwan) according to the manufacturer's recommendations (Hu et al., 2022).…”

Section: Methodsmentioning

confidence: 99%

“…SNaPshot/high‐throughput ligation‐dependent probe amplification (HLPA) was applied to detect 61 known mutational hotspots of the α‐ and β‐globin genes (Table 1). The experiment was conducted as previously described (Hu et al., 2022; Shao et al., 2021). HBA1/2 (NM_000558.3/NM_000517.4) and HBB (NM_000518.4) point mutations were genotyped by using single‐nucleotide polymorphism genotyping with the multiplex ligation detection reaction technique.…”

Section: Methodsmentioning

confidence: 99%

“…As a mature genotyping technology, capillary electrophoresis has been widely used in DNA sequencing analysis owing to its short run time and ease of operation (Hu et al., 2022; Shao et al., 2021). In the present study, we developed a capillary electrophoresis‐based multiplex PCR assay allowing the simultaneous detection of the most common pathogenic variants among HBA1/HBA2 and HBB genes in the Chinese population.…”

Section: Introductionmentioning

confidence: 99%

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Mutation spectrum of thalassemia among pre‐pregnant adults in the Jiangsu Province by capillary electrophoresis‐based multiplex PCR assay

Shao,

Wang,

Zhang

et al. 2023

Molec Gen & Gen Med

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BackgroundThalassemia is a common genetic disorder in southwestern China, and an increasing number of cases from eastern China have been recently reported. Here, we developed a rapid, convenient, and accurate assay to evaluate the mutation spectrum of thalassemia in eastern China.MethodsA carrier screening assay for 61 hotspot variants among HBA1/HBA2 and HBB (OMIM: 141800, 141850, and 141900) genes was developed by SNaPshot/high‐throughput ligation‐dependent probe amplification (HLPA) technology. We used this assay to detect the mutation spectrum of thalassemia in individuals from eastern China and compared with the data collected from literatures focused on southern and northern China for variant distribution.ResultsAmong 4276 tested individuals, 2.62% (112/4276) were α‐thalassemia carriers, with 90 carrying one deletion or mutation and 22 carrying two deletions. 0.40% (17/4276) were β‐thalassemia carriers, and the most common variant of β‐thalassemia was c.126_129delCTTT (29.41%) followed by c.316‐197C>T (23.53%). The genotype distribution in our study was similar to those from southern China populations.ConclusionThe Chinese population from different regions presented comparable mutation spectrum of thalassemia, and the SNaPshot/HLPA technique may serve as a capable assay for a routine genetic test in clinical practice with its accurate, rapid, and inexpensive advantage.

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Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Mutation spectrum of thalassemia among pre‐pregnant adults in the Jiangsu Province by capillary electrophoresis‐based multiplex PCR assay

Shao,

Wang,

Zhang

et al. 2023

Molec Gen & Gen Med

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“…Pedigree 1, Index patient (II 4), a 37-year-old female, planned to undergo preimplantation genetic testing for structural rearrangement (PGT-SR) because of carrying the Robertson translocation chromosome. Before PGT-SR, the couple voluntarily selected expanded carrier screening (ECS) for recessive genetic diseases using a previously reported method [19], which suggested the female patient carried a duplication variant of exons 1-2 in the canonical isoform of DMD gene. To analyze the pathogenicity of the variant and evaluate the reproductive risk, the family investigation was performed, and no members with DMD/BMD-related phenotypes were found.…”

Section: Subjectsmentioning

confidence: 99%

Comprehensive analysis of genomic complexity in the 5’ end coding region of the DMD gene in patients of exons 1-2 duplications based on long-read sequencing

Shen,

Ding,

Sun

et al. 2023

Preprint

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Backgroud Dystrophinopathies are the most common X-linked inherited muscle diseases, and the disease-causing gene is DMD. Exonic duplications are common type of pathogenic variants in DMD gene, however, 5’ end exonic duplications containing exon 1 are less common. When assessing the pathogenicity of exonic duplications in the DMD gene, consideration must be given to their impact on the reading frame. Traditional molecular methods, such as multiplex ligation-dependent probe amplification (MLPA) and next generation sequencing (NGS), are commonly used in clinic. However, they cannot discriminate the precise physical locations of breakpoints and structural features of genomic rearrangement. Long-read sequencing (LRS) can effectively overcome this limitation. Results We report three cases involving the duplications of exon 1 and/or exon 2 in DMD gene. The locations of the breakpoints and the characteristics of structural variants (SVs) are identified through LRS, enabling the classification of the variants' pathogenicity. Two distinct variant types encompassing exon 1 in the DMD Dp427m isoform and/or Dp427c isoform are identified, which have been infrequently reported previously. The male individuals harboring duplication variant of consecutive exons 1–2 in the DMD canonical transcript (Dp427m) and exon 1 in the Dp427c transcript are normal, indicating the variant is benign. However, one patient carries a complex SVs involving exon 1 of the DMD Dp427c transcript showing an obvious phenotype. Conclusions Our research sheds light on the complexity of DMD variants encompassing Dp427c/Dp427m promoter regions and emphasizes the importance of cautious interpretation when assessing the pathogenicity of DMD 5' end exonic duplications, particularly in carrier screening scenarios without an affected proband.

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Comprehensive analysis of genomic complexity in the 5’ end coding region of the DMD gene in patients of exons 1–2 duplications based on long-read sequencing

Shen,

Ding,

Sun

et al. 2024

BMC Genomics

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Background Dystrophinopathies are the most common X-linked inherited muscle diseases, and the disease-causing gene is DMD. Exonic duplications are a common type of pathogenic variants in the DMD gene, however, 5’ end exonic duplications containing exon 1 are less common. When assessing the pathogenicity of exonic duplications in the DMD gene, consideration must be given to their impact on the reading frame. Traditional molecular methods, such as multiplex ligation-dependent probe amplification (MLPA) and next-generation sequencing (NGS), are commonly used in clinics. However, they cannot discriminate the precise physical locations of breakpoints and structural features of genomic rearrangement. Long-read sequencing (LRS) can effectively overcome this limitation. Results We used LRS technology to perform whole genome sequencing on three families and analyze the structural variations of the DMD gene, which involves the duplications of exon 1 and/or exon 2. Two distinct variant types encompassing exon 1 in the DMD Dp427m isoform and/or Dp427c isoform are identified, which have been infrequently reported previously. In pedigree 1, the male individuals harboring duplication variant of consecutive exons 1–2 in the DMD canonical transcript (Dp427m) and exon 1 in the Dp427c transcript are normal, indicating the variant is likely benign. In pedigree 3, the patient carries complex SVs involving exon 1 of the DMD Dp427c transcript showing an obvious phenotype. The locations of the breakpoints and the characteristics of structural variants (SVs) are identified by LRS, enabling the classification of the variants' pathogenicity. Conclusions Our research sheds light on the complexity of DMD variants encompassing Dp427c/Dp427m promoter regions and emphasizes the importance of cautious interpretation when assessing the pathogenicity of DMD 5' end exonic duplications, particularly in carrier screening scenarios without an affected proband.

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A capillary electrophoresis-based multiplex PCR assay for expanded carrier screening in the eastern Han Chinese population

Cited by 4 publications

References 55 publications

Mutation spectrum of thalassemia among pre‐pregnant adults in the Jiangsu Province by capillary electrophoresis‐based multiplex PCR assay

Mutation spectrum of thalassemia among pre‐pregnant adults in the Jiangsu Province by capillary electrophoresis‐based multiplex PCR assay

Comprehensive analysis of genomic complexity in the 5’ end coding region of the DMD gene in patients of exons 1-2 duplications based on long-read sequencing

Comprehensive analysis of genomic complexity in the 5’ end coding region of the DMD gene in patients of exons 1–2 duplications based on long-read sequencing

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