Single-cell Genome-Wide Concurrent Haplotyping and Copy-Number Profiling Through Genotyping-by-Sequencing
Summary
Leveraging single-cell whole-genome haplotyping facilitates simultaneous detection of haplotypes linked to monogenic diseases, chromosomal copy-numbering, and the unearthing of mosaicism in embryos and embryonic stem cells. Utilizing karyomapping and haplarithmisis as generic, genome-wide modalities, these techniques supplant conventional preimplantation genetic testing (PGT). Although single-nucleotide polymorphism (SNP) arrays predominate, sequencing-based methodologies are predicted to gain accessibility and affordability. The researchers crafted an innovative sequencing-based approach for single-cell haplotyping and copy-number profiling. Following DNA amplification and restriction enzyme digestion, genotyping occurs through sequencing. This single-cell genotyping-by-sequencing (scGBS) serves as input for haplarithmisis, enabling accurate concurrent haplotyping and copy-number profiling of single cells. Demonstrating applicability in human blastomere and trophectoderm samples for PGT of monogenic disorders, the method also proved effective in bovine embryo blastomeres, expanding potential clinical PGT applications across diploid species.
Research Criteria
The authors devise an innovative sequencing-based technique for concurrent haplotyping and copy-number profiling in single cells. Employing DNA amplification, restriction enzyme digestion, and sequencing, they introduce single-cell genotyping-by-sequencing (scGBS) as an input for haplarithmisis—an algorithm established for SNP array-based single-cell haplotyping. The researchers formulated technical parameters and crafted an analytical pipeline, which substantiates precise haplotyping and copy-number analysis.
Fig.1 Experimental design. (Masset, 2022)
Sample Type
Human blastomere and trophectoderm samples, bovine embryos, and human cell lines.
Result—scGBS on HapMap Cell Lines
The article delineates the development of a novel sequencing-based method for single-cell haplotyping and copy-number profiling, utilizing DNA amplification, restriction enzyme digestion, and sequencing to establish single-cell genotyping-by-sequencing (scGBS) as an input for haplarithmisis. The authors crafted a precise analytical pipeline for concurrent haplotyping and copy-number profiling, demonstrating its utility in human blastomere and trophectoderm samples for preimplantation genetic testing of monogenic disorders, and applicability in other species. The scGBS approach was tested on HapMap cell lines, revealing optimal depths of coverage for bulk and single-cell sequencing. The resultant scGBS haplotypes demonstrated high concordance with both SNP array-derived and bulk scGBS haplotypes, establishing the method's accuracy and reliability.
Fig.2 scGBS haplotyping and copy-number profiling on HapMap cell lines. (Masset, 2022)
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Reference
- Masset, H.; et al. Single-cell genome-wide concurrent haplotyping and copy-number profiling through genotyping-by-sequencing. Nucleic Acids Research. 2022, 50(11): e63.
